Acupoint Sensitization, Acupuncture Analgesia, Acupucnture On Visceral Functional Disorders and Its Mechanism

Evidence-Based Complementary and Alternative Medicine
Acupoint Sensitization,
Acupuncture Analgesia,
Acupuncture on Visceral
Functional Disorders,
and Its Mechanism
Guest Editors: Xiaochun Yu, Bing Zhu, Zhixiu Lin, Haifa Qiao, Jian Kong,
and Xinyan Gao
Acupoint Sensitization, Acupuncture Analgesia,
Acupuncture on Visceral Functional Disorders,
and Its Mechanism
Acupoint Sensitization, Acupuncture Analgesia,

Acupuncture on Visceral Functional Disorders,
and Its Mechanism
Guest Editors: Xiaochun Yu, Bing Zhu, Zhixiu Lin, Haifa Qiao,
Jian Kong, and Xinyan Gao
Copyright © 2015 Hindawi Publishing Corporation. All rights reserved.
This is a special issue published in “Evidence-Based Complementary and Alternative Medicine .” All articles are open access articles
distributed under the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any
medium, provided the original work is properly cited.
Editorial Board
Mona Abdel-Tawab, Germany Shun-Wan Chan, Hong Kong Filippo Fratini, Italy
Jon Adams, Australia Il-Moo Chang, Republic of Korea Brett Froeliger, USA
Gabriel A. Agbor, Cameroon Chun T. Che, USA Maria pia Fuggetta, Italy
Ulysses P. Albuquerque, Brazil Kevin Chen, USA Joel J. Gagnier, Canada
S. L. Aleryani, USA Evan P. Cherniack, USA Siew Hua Gan, Malaysia
Ather Ali, USA Salvatore Chirumbolo, Italy Jian-Li Gao, China
M. S. Ali-Shtayeh, State of Palestine Jae Youl Cho, Republic of Korea Mary K. Garcia, USA
Gianni Allais, Italy K. B. Christensen, Denmark Susana Garcia de Arriba, Germany
Terje Alraek, Norway Shuang-En Chuang, Taiwan D. G. Giménez, Spain
Shrikant Anant, USA Y. N. Clement, Trinidad and Tobago Gabino Garrido, Chile
Isabel Andjar, Spain Paolo Coghi, Italy Ipek Goktepe, Qatar
Letizia Angiolella, Italy Marisa Colone, Italy Michael Goldstein, USA
Virginia A. Aparicio, Spain Lisa A. Conboy, USA Yuewen Gong, Canada
Makoto Arai, Japan Kieran Cooley, Canada Settimio Grimaldi, Italy
Hyunsu Bae, Republic of Korea Edwin L. Cooper, USA Gloria Gronowicz, USA
Giacinto Bagetta, Italy Olivia Corcoran, UK Maruti Ram Gudavalli, USA
Onesmo B. Balemba, USA Muriel Cuendet, Switzerland Alessandra Guerrini, Italy
Winfried Banzer, Germany Roberto K. N. Cuman, Brazil Narcis Gusi, Spain
Panos Barlas, UK Vincenzo De Feo, Italy Svein Haavik, Norway
Vernon A. Barnes, USA Rocı́o De la Puerta, Spain Solomon Habtemariam, UK
Samra Bashir, Pakistan Laura De Martino, Italy Abid Hamid, India
Purusotam Basnet, Norway Nunziatina De Tommasi, Italy Michael G. Hammes, Germany
Jairo Kennup Bastos, Brazil Alexandra Deters, Germany Kuzhuvelil B. Harikumar, India
Sujit Basu, USA Farzad Deyhim, USA Cory S. Harris, Canada
Arpita Basu, USA Manuela Di Franco, Italy Jan Hartvigsen, Denmark
George David Baxter, New Zealand Claudia Di Giacomo, Italy Thierry Hennebelle, France
André-Michael Beer, Germany Antonella Di Sotto, Italy Lise Hestbaek, Denmark
Alvin J. Beitz, USA M.-G. Dijoux-Franca, France Eleanor Holroyd, Australia
Louise Bennett, Australia Luciana Dini, Italy Markus Horneber, Germany
Maria Camilla Bergonzi, Italy Tieraona L. Dog, USA Ching-Liang Hsieh, Taiwan
Anna R. Bilia, Italy Caigan Du, Canada Benny T. K. Huat, Singapore
Yong C. Boo, Republic of Korea Jeng-Ren Duann, Taiwan Roman Huber, Germany
Monica Borgatti, Italy Nativ Dudai, Israel Helmut Hugel, Australia
Francesca Borrelli, Italy Thomas Efferth, Germany Ciara Hughes, UK
Gloria Brusotti, Italy Abir El-Alfy, USA Attila Hunyadi, Hungary
Arndt Büssing, Germany Tobias Esch, USA Sumiko Hyuga, Japan
Rainer W. Bussmann, USA Giuseppe Esposito, Italy H. Stephen Injeyan, Canada
Andrew J. Butler, USA Keturah R. Faurot, USA Chie Ishikawa, Japan
Gioacchino Calapai, Italy Yibin Feng, Hong Kong Angelo A. Izzo, Italy
Giuseppe Caminiti, Italy Nianping Feng, China Chris J. Branford-White, UK
Raffaele Capasso, Italy Patricia D. Fernandes, Brazil Suresh Jadhav, India
Francesco Cardini, Italy Josue Fernandez-Carnero, Spain G. K. Jayaprakasha, USA
Opher Caspi, Israel Antonella Fioravanti, Italy Stefanie Joos, Germany
Subrata Chakrabarti, Canada Fabio Firenzuoli, Italy Zeev L Kain, USA
Pierre Champy, France Peter Fisher, UK Osamu Kanauchi, Japan
Wenyi Kang, China Karin Meissner, Germany Cheppail Ramachandran, USA
Shao-Hsuan Kao, Taiwan Albert S Mellick, Australia Elia Ranzato, Italy
Juntra Karbwang, USA A. G. Mensah-Nyagan, France Ke Ren, USA
Kenji Kawakita, Japan Andreas Michalsen, Germany Man H. Rhee, Republic of Korea
Deborah A. Kennedy, Canada Oliver Micke, Germany Luigi Ricciardiello, Italy
C.-H. Kim, Republic of Korea Roberto Miniero, Italy Daniela Rigano, Italy
Y. C. Kim, Republic of Korea Giovanni Mirabella, Italy José L. Rı́os, Spain
Yoshiyuki Kimura, Japan David Mischoulon, USA Paolo Roberti di Sarsina, Italy
Toshiaki Kogure, Japan Francesca Mondello, Italy Mariangela Rondanelli, Italy
Jian Kong, USA Albert Moraska, USA Omar Said, Israel
Tetsuya Konishi, Japan Giuseppe Morgia, Italy Avni Sali, Australia
Karin Kraft, Germany Mark Moss, UK Mohd Z. Salleh, Malaysia
Omer Kucuk, USA Yoshiharu Motoo, Japan Andreas Sandner-Kiesling, Austria
Victor Kuete, Cameroon Kamal D. Moudgil, USA Manel Santafe, Spain
Yiu W. Kwan, Hong Kong Yoshiki Mukudai, Japan Tadaaki Satou, Japan
Kuang C. Lai, Taiwan Frauke Musial, Germany Michael A. Savka, USA
Ilaria Lampronti, Italy MinKyun Na, Republic of Korea Claudia Scherr, Switzerland
Lixing Lao, Hong Kong Hajime Nakae, Japan G. Schmeda-Hirschmann, Chile
Christian Lehmann, Canada Srinivas Nammi, Australia Andrew Scholey, Australia
Marco Leonti, Italy Krishnadas Nandakumar, India Roland Schoop, Switzerland
Lawrence Leung, Canada Vitaly Napadow, USA Sven Schröder, Germany
Shahar Lev-ari, Israel Michele Navarra, Italy Herbert Schwabl, Switzerland
Xiu-Min Li, USA Isabella Neri, Italy Veronique Seidel, UK
Chun G. Li, Australia Pratibha V. Nerurkar, USA Senthamil R. Selvan, USA
Min Li, China Karen Nieber, Germany Felice Senatore, Italy
Bi-Fong Lin, Taiwan Menachem Oberbaum, Israel Hongcai Shang, China
Ho Lin, Taiwan Martin Offenbaecher, Germany Ronald Sherman, USA
Christopher G. Lis, USA Junetsu Ogasawara, Japan Karen J. Sherman, USA
Gerhard Litscher, Austria Ki-Wan Oh, Republic of Korea Kuniyoshi Shimizu, Japan
I-Min Liu, Taiwan Yoshiji Ohta, Japan Kan Shimpo, Japan
Yijun Liu, USA Olumayokun A. Olajide, UK Yukihiro Shoyama, Japan
Vı́ctor López, Spain Thomas Ostermann, Germany Morry Silberstein, Australia
Thomas Lundeberg, Sweden Siyaram Pandey, Canada K. N. S. Sirajudeen, Malaysia
Filippo Maggi, Italy Bhushan Patwardhan, India Graeme Smith, UK
Valentina Maggini, Italy Berit S. Paulsen, Norway Chang-Gue Son, Republic of Korea
Gail B. Mahady, USA Philip Peplow, New Zealand Rachid Soulimani, France
Jamal Mahajna, Israel Florian Pfab, Germany Didier Stien, France
Juraj Majtan, Slovakia Sonia Piacente, Italy Con Stough, Australia
Francesca Mancianti, Italy Andrea Pieroni, Italy Annarita Stringaro, Italy
Carmen Mannucci, Italy Richard Pietras, USA Shan-Yu Su, Taiwan
A.-M. Manuel, Spain Andrew Pipingas, Australia Barbara Swanson, USA
Fulvio Marzatico, Italy Jose M. Prieto, UK Giuseppe Tagarelli, Italy
Marta Marzotto, Italy Haifa Qiao, USA O. Taglialatela-Scafati, Italy
James H. McAuley, Australia Waris Qidwai, Pakistan Takashi Takeda, Japan
Kristine McGrath, Australia Xianqin Qu, Australia Ghee T. Tan, USA
James S. McLay, UK E. Ferreira Queiroz, Switzerland Hirofumi Tanaka, USA
Lewis Mehl-Madrona, USA Roja Rahimi, Iran Lay Kek Teh, Malaysia
Peter Meiser, Germany Khalid Rahman, UK Norman Temple, Canada
Mayank Thakur, Germany Giuseppe Venturella, Italy Darren R. Williams, Republic of Korea
Menaka C. Thounaojam, USA Pradeep Visen, Canada Christopher Worsnop, Australia
Evelin Tiralongo, Australia Aristo Vojdani, USA Haruki Yamada, Japan
Stephanie Tjen-A-Looi, USA Dawn Wallerstedt, USA Nobuo Yamaguchi, Japan
MichaThl Tomczyk, Poland Shu-Ming Wang, USA Ling Yang, China
Loren Toussaint, USA Chong-Zhi Wang, USA Junqing Yang, China
Yew-Min Tzeng, Taiwan Yong Wang, USA Eun J. Yang, Republic of Korea
Dawn M. Upchurch, USA Jonathan L. Wardle, Australia Ken Yasukawa, Japan
Konrad Urech, Switzerland Kenji Watanabe, Japan Albert S. Yeung, USA
Takuhiro Uto, Japan J. Wattanathorn, Thailand Armando Zarrelli, Italy
Sandy van Vuuren, South Africa Michael Weber, Germany Chris Zaslawski, Australia
Alfredo Vannacci, Italy Silvia Wein, Germany Ruixin Zhang, USA
Subramanyam Vemulpad, Australia Janelle Wheat, Australia
Carlo Ventura, Italy Jenny M. Wilkinson, Australia
Contents
Acupoint Sensitization, Acupuncture Analgesia, Acupuncture on Visceral Functional Disorders, and Its
Mechanism, Xiaochun Yu, Bing Zhu, Zhixiu Lin, Haifa Qiao, Jian Kong, and Xinyan Gao
Volume 2015, Article ID 171759, 1 pages
Influences of Deqi on Immediate Analgesia Effect of Needling SP6 (Sanyinjiao) in Patients with
Primary Dysmenorrhea in Cold and Dampness Stagnation Pattern: Study Protocol for a Randomized
Controlled Trial, Yu-qi Liu, Peng Zhang, Jie-ping Xie, Liang-xiao Ma, Hong-wen Yuan, Jing Li, Chi Lin,
Pei Wang, Guo-yan Yang, and Jiang Zhu
Acupuncture for Functional Dyspepsia: A Single Blinded, Randomized, Controlled Trial, Yulian Jin,
Qing Zhao, Kehua Zhou, Xianghong Jing, Xiaochun Yu, Jiliang Fang, Zhishun Liu, and Bing Zhu
Discovery of Acupoints and Combinations with Potential to Treat Vascular Dementia: A Data Mining
Analysis, Shuwei Feng, Yulan Ren, Shilin Fan, Minyu Wang, Tianxiao Sun, Fang Zeng, Ping Li, and Fanrong
Liang
Effects of Deep Electroacupuncture Stimulation at “Huantiao” (GB 30) on Expression of

Apoptosis-Related Factors in Rats with Acute Sciatic Nerve Injury, Lili Dai, Yanjing Han, Tieming Ma,
Yuli Liu, Lu Ren, Zenghua Bai, and Ye Li
Function of Nucleus Ventralis Posterior Lateralis Thalami in Acupoint Sensitization Phenomena,

Pei-Jing Rong, Jing-Jun Zhao, Ling-Ling Yu, Liang Li, Hui Ben, Shao-Yuan Li, and Bing Zhu
Analysis and Thoughts about the Negative Results of International Clinical Trials on Acupuncture,
Wei-hong Liu, Yang Hao, Yan-jing Han, Xiao-hong Wang, Chen Li, and Wan-ning Liu
Efficacy of Acupuncture in Children with Nocturnal Enuresis: A Systematic Review and Meta-Analysis
of Randomized Controlled Trials, Zheng-tao Lv, Wen Song, Jing Wu, Jun Yang, Tao Wang, Cai-hua Wu,
Fang Gao, Xiao-cui Yuan, Ji-hong Liu, and Man Li
Regulation of Neurotrophin-3 and Interleukin-1𝛽 and Inhibition of Spinal Glial Activation Contribute
to the Analgesic Effect of Electroacupuncture in Chronic Neuropathic Pain States of Rats, Wenzhan Tu,
Wansheng Wang, Haiyan Xi, Rong He, Liping Gao, and Songhe Jiang
Brain Network Response to Acupuncture Stimuli in Experimental Acute Low Back Pain: An fMRI
Study, Yu Shi, Ziping Liu, Shanshan Zhang, Qiang Li, Shigui Guo, Jiangming Yang, and Wen Wu
Effect of Repeated Electroacupuncture Intervention on Hippocampal ERK and p38MAPK Signaling in

Neuropathic Pain Rats, Jun-ying Wang, Shu-ping Chen, Yong-hui Gao, Li-na Qiao, Jian-liang Zhang,
and Jun-ling Liu
Eye Acupuncture Treatment for Stroke: A Systematic Review and Meta-Analysis, Zeng-Hua Bai,
Zhi-Xing Zhang, Chun-Ri Li, Mei Wang, Meong-Ju Kim, Hui Guo, Chun-Yan Wang, Tong-Wu Xiao,
Yuan Chi, Lu Ren, Zhong-Yue Gu, and Ran Xu
Effect of Acupuncture on Functional Connectivity of Anterior Cingulate Cortex for Bell’s Palsy Patients
with Different Clinical Duration, Hongli Wu, Hongxing Kan, Chuanfu Li, Kyungmo Park, Yifang Zhu,
Abdalla Z. Mohamed, Chunsheng Xu, Yuanyuan Wu, Wei Zhang, and Jun Yang
The Study of Dynamic Characteristic of Acupoints Based on the Primary Dysmenorrhea Patients with
the Tenderness Reflection on Diji (SP 8), Sheng Chen, Yanhuan Miao, Yinan Nan, Yanping Wang, Qi Zhao,
Enhui He, Yini Sun, and Jiping Zhao
Hindawi Publishing Corporation
Volume 2015, Article ID 171759, 1 page
http://dx.doi.org/10.1155/2015/171759
Editorial
Acupoint Sensitization, Acupuncture Analgesia, Acupuncture on
Visceral Functional Disorders, and Its Mechanism
Xiaochun Yu,1 Bing Zhu,1 Zhixiu Lin,2 Haifa Qiao,3 Jian Kong,4 and Xinyan Gao1
1
Institute of Acupuncture, China Academy of Chinese Medical Sciences, 16 Nanxiaojie, Dongzhimen, Beijing 100700, China
2
School of Chinese Medicine, Faculty of Medicine, Chinese University of Hong Kong, Shatin, New Territories, Hong Kong
3
Department of Biomedical Sciences, College of Medicine, Florida State University, 1115 W. Call Street, Tallahassee, FL 32306, USA
4
Massachusetts General Hospital, Building 120, 2nd Street, Suite 101C, Charlestown, MA, USA
Correspondence should be addressed to Xiaochun Yu; yuxc@mail.cintcm.ac.cn
Received 14 May 2015; Accepted 21 May 2015
Copyright © 2015 Xiaochun Yu et al. This is an open access article distributed under the Creative Commons Attribution License,
which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited.
Our special issue, which had opened for 6 months in blinded, randomized, controlled trial on acupuncture treat-
the second half of 2014, focused on acupoint sensitization, ment of functional dyspepsia and found that acupuncture
acupuncture analgesia, acupuncture for visceral modulation manipulation exhibited better effects on improving dyspeptic
in gastrointestinal systems, acupuncture for modulation of symptoms, mental status, and quality of life in patients with
brain function, acupoint combination treatment of insomnia FD than nonacupoint without manipulation.
and gastrointestinal disorders, and nonspecific and specific In general, we have papers involving clinical trials, data
effects of acupuncture based on stimulation intensity. mining analysis or study protocol, and basic research in press,
Of these papers in press, S. Chen et al. reported that which thoroughly meet the expectation of our initial call for
the location and tenderness of Diji (SP8) were not the same papers of this issue.
in healthy subjects as in dysmenorrheal patients, suggesting
dynamic and sensitivity of acupoints under different patho- Acknowledgments
logical status. S. Feng et al. did data mining analysis on acu-
points or combinations for treatment of vascular dementia We thank all authors for their excellent contributions and
and gave suggestions for acupoint selection based on the reviewers for their valuable help. The Lead Guest Editor
most commonly used formulas. L. Dai et al. performed basic would like to thank the five Guest Editors for their dedicated
research in a sciatic nerve injury rat model and found that cooperation. We hope the special issue will bring readers
deep EA stimulation is better in improving neuromuscular useful academic reference in their research.
function and benign regulation of apoptosis-related factors
than shallow EA. J. Wang et al., based on their previous Xiaochun Yu
study that hippocampal mAChR-1 participating in MARK Bing Zhu
signaling was involved in EA induced cumulative analgesia Zhixiu Lin
in neuropathic pain rats, observed in their present study that Haifa Qiao
EA2W was closely related to the cumulative analgesia via Jian Kong
intracellular ERK and p38 MARK signaling. P. Rong et al. Xinyan Gao
observed that, in anesthetized rats, EA on ST36-ST37 could
enhance nucleus ventralis posterior lateralis thalami neu-
ronal discharges which were fired by CRD-induced visceral
pain. Their study indicates that acupoints may be sensitized
under visceral disorders. Y. Jin et al. conducted a single
http://dx.doi.org/10.1155/2015/238790
Research Article
Influences of Deqi on Immediate Analgesia Effect of
Needling SP6 (Sanyinjiao) in Patients with Primary
Dysmenorrhea in Cold and Dampness Stagnation Pattern:
Study Protocol for a Randomized Controlled Trial
Yu-qi Liu,1,2 Peng Zhang,1,3 Jie-ping Xie,1,4 Liang-xiao Ma,1,4 Hong-wen Yuan,5
Jing Li,1 Chi Lin,1 Pei Wang,1 Guo-yan Yang,1 and Jiang Zhu1,4
1
School of Acupuncture Moxibustion and Tuina, Beijing University of Chinese Medicine, Beijing 100029, China
2
Institute of Basic Research in Clinical Medicine, China Academy of Chinese Medical Sciences, Beijing 100700, China
3
Beijing Tongren Hospital Affiliated to Capital Medical University, Beijing 100730, China
4
The Key Unit of State Administration of Traditional Chinese Medicine, Evaluation of Characteristic Acupuncture Therapy,
Beijing 100029, China
5
School of Traditional Chinese Medicine, Capital Medical University, Beijing 100069, China
Correspondence should be addressed to Jiang Zhu; jzhjzh@263.net
Received 22 August 2014; Revised 9 November 2014; Accepted 19 November 2014
Academic Editor: Xinyan Gao
Copyright © 2015 Yu-qi Liu et al. This is an open access article distributed under the Creative Commons Attribution License, which
permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited.
Deqi, according to traditional Chinese medicine, is a specific needle sensation during the retention of needles at certain acupoints
and is considered to be necessary to produce therapeutic effects from acupuncture. Although some modern researches have showed
that Deqi is essential for producing acupuncture analgesia and anesthesia, the data are not enough. It is a paper of a multicenter,
randomized controlled study protocol, to evaluate the influences of Deqi on acupuncture SP6 in Cold and Dampness Stagnation
pattern primary dysmenorrhea patients, in terms of reducing pain and anxiety, and to find out the relationship between Deqi and
the temperature changes at SP6 (Sanyinjiao) and CV4 (Guanyuan). The results of this trial will be helpful to explain the role of Deqi
in acupuncture analgesia and may provide a new objective index for measuring Deqi in the future study. This trial is registered with
ChiCTR-TRC-13003086.
1. Introduction the response of brain [5, 6]. For acupuncturists, they can feel a
change of the mechanical behavior of the tissues surrounding
Deqi, according to records in both ancient and modern the needle, such as an increase of the force necessary to pull
books of traditional Chinese medicine (TCM), is a specific the needle out of the tissue (pullout force) [7]. Although some
needle sensation which usually occurs during retention of modern research showed that Deqi is essential for producing
acupuncture needles at certain acupoints, and both patients acupuncture analgesia and anesthesia [8], new evidence to
and acupuncturists can feel the Deqi sensation. It is regarded confirm the conclusion is still urgently needed [9, 10].
as a necessary factor for producing effects from acupuncture Primary dysmenorrhea (PD) refers to painful menstrual
following traditional Chinese medicine theory. Patients can cramps without any evident pathology. It is characterized
feel Deqi as multiple sensations at the needling acupoints and by crampy suprapubic pain with radiation into the lower
along the meridians sometime, such as soreness, numbness, quadrants, the lumbar area, and the thighs [11]. PD is a com-
distension, or a minimal muscular contraction around the mon gynecological complaint and significantly affects study,
needle [1–3] and also objective physiological changes, such work, sports, and social activities [12–18]. So far nonsteroidal
as the skin temperature changes at the acupoints [4] or anti-inflammatory drugs (NSAIDs) or oral contraceptive
2 Evidence-Based Complementary and Alternative Medicine
pills (OCPs) are widely advocated as standard treatments calculating sample size for further studies. Assuming a
for women with PD [19, 20]. However, acupuncture as dropout rate of 20%, we will plan to enroll a total of 96
nonpharmacological approaches has great potential value. participants with 48 in each group.
Acupuncture is one of the main treatment modalities of
TCM. Several trials [21, 22] have already demonstrated the 2.2.2. Recruitment of Participants. Participants will be
encouraging results of acupuncture as a nonpharmacological recruited from the four hospitals mentioned above in outpa-
option for the treatment of PD. Two systematic reviews also tient clinics. Posters will be used outside the acupuncture
demonstrated the effect [23] and cost-effectiveness [24] of clinics. The posters will contain brief introductions about the
this therapy. The acupoint SP6 (Sanyinjiao) is found to be one inclusion/exclusion criteria, the free acupuncture treatments
of the most commonly used points encountered when search- offered to eligible participants, and the contact information
ing ancient Chinese medical classics, Chinese acupuncture of the researchers.
textbooks, and clinical trials using acupuncture-related ther-
apies for PD [25, 26]. Previous randomized controlled trials 2.2.3. Inclusion Criteria. Participants who meet all the follow-
have showed that acupuncture at SP6 (Sanyinjiao) can relieve ing requirements will be allowed for enrollment:
the pain of PD immediately [27–29] especially for PD patients
with Cold and Dampness Stagnation pattern [30], which is (1) nulliparous women, aged between 18 and 30 years old,
the most common pattern in PD patients [31, 32]. Langevin’s diagnosed with PD (according to the criteria of the
study [7] also showed that, compared with the nonacupoint Primary Dysmenorrhea Consensus Guideline [34]),
control group, the average pullout force in SP6 (Sanyinjiao) and in Cold and Dampness Stagnation pattern (based
was greater, which suggested that SP6 (Sanyinjiao) might be on a revised Chinese national guideline [30]),
especially sensitive in Deqi. (2) moderate to severe primary dysmenorrhea (PD)
In this study, we will perform a randomized controlled (≥40 mm on VAS-P) for three consecutive menstrual
trial using SP6 to investigate the influences of Deqi on cycles,
immediate analgesia effects in PD patients with Cold and
(3) duration of PD (self-reported pain) varying from 6
Dampness Stagnation pattern. The primary objective of this
months to 15 years,
trial is to evaluate the influences of Deqi on acupuncture
therapy, in terms of pain reduction (measured on a 0–100 mm (4) written informed consent.
visual analogue scale for pain (VAS-P)) achieved before
and after intervention (i.e., before and after 30 minutes of 2.2.4. Exclusion Criteria. Exclusion criteria are as follows:
treatment). The secondary objectives are as follows: (1) to
evaluate the influences of anxiety reduction on a 0–100 mm (1) secondary dysmenorrhea (e.g., endometriosis and
visual analogue scale for anxiety (VAS-A) and (2) to evaluate fibroids),
the influences of temperature changes at SP6 (Sanyinjiao) (2) irregular/infrequent menstrual cycles (i.e., beyond
and CV4 (Guanyuan) acupoints monitor by a digital infrared the typical range of 21- to 35-day cycle),
thermographic imaging device (only in the Dongzhimen
(3) complication with severe diseases (e.g., cerebral, liver,
Hospital).
kidney, or hematopoietic system diseases), mental
defects, or asthma,
2. Methods
(4) pregnancy,
2.1. Ethics. The trial protocol is in accordance with the (5) use of analgesic medication in 24 hours before treat-
principle of the Declaration of Helsinki [33] and has been ment,
approved by the Ethic Committee of Beijing University of
Chinese Medicine (Beijing, China, Approval number 2012- (6) having a professional acupuncture background,
040). Each participant will be notified regarding the study (7) having potentially poor treatment compliance (e.g.,
protocol. Written informed consent will be obtained from unstable working and living situation or difficulty in
each participant. following up).
2.2. Settings and Participants. A target sample of 96 par- 2.3. Randomization and Blinding. The central randomization
ticipants will be recruited in acupuncture clinic from the will be performed by the Center for Evidence-Based Chinese
following four hospitals: Dongzhimen Hospital Affiliated to Medicine affiliated to Beijing University of Chinese Medicine
Beijing University of Chinese Medicine, Beijing Hospital in China, using complete randomization to generate the
of Traditional Chinese Medicine affiliated to Capital Med- random allocation sequence. Once a participant is included,
ical University, Huguosi Hospital of Traditional Chinese the researcher will contact the randomization center for the
Medicine affiliated to Beijing University of Chinese Medicine, group allocation of the participant, and the acupuncturist will
and Hebei Medical University. The trial will be conducted be informed by telephone. Participants will be allocated at a
from March 2013 to March 2015. 1 : 1 ratio (Figure 1).
The participants will be informed that they will have a
2.2.1. Sample Size. This is a study without similar references 50% chance of being allocated in either of the two acupunc-
as we know and 40 patients per group is acceptable for ture groups, and both groups will be potentially effective;
Evidence-Based Complementary and Alternative Medicine 3
Participants with PD
Excluded
- Not meeting
Check inclusion criteria inclusion criteria
and offer the study
- Meeting exclusion
criteria
- Refusing to
Agree to participate
participate
Randomization
1:1
Allocated to Deqi Allocated to no Deqi
One treatment for 30 minutes
Evaluation of results at the end of treatment
Figure 1: Flow diagram for the study. Work scheme with description of assessment visits and times.
hence, participants will be blinded to the group allocation. 2.5. Outcome Measures
The patients will also be informed of the possible risks
2.5.1. Primary Outcome Measures. The changes of pain will be
associated with acupuncture (hematoma or fainting). The
measured by visual analogue scale for pain (VAS-P), before
acupuncturist cannot be blinded due to the specific nature
and after the treatment. The validity and reliability of the
of intervention. Outcome assessors and personnel who will
VAS-P scale have been proven [36–38] and it was employed
deal with data collection and data analysis will be blinded
in our previous similar studies [29, 30]. The scale measures
throughout the entire trial.
a continuous quantitative variable varying from 0 mm (no
pain) to 100 mm (worst pain ever).
2.4. Interventions. All participants, when their VAS-P score
of menstrual pain is equal to or more than 40 mm on the
first day of menstruation, will each receive an acupuncture 2.5.2. Secondary Outcome Measures
treatment for 30 minutes at bilateral SP6 acupoints. The SP6 (1) The changes of the anxiety are measured by visual
lies on the tibial aspect of the leg, is posterior to the medial analogue scale for anxiety (VAS-A), before and after
border of the tibia, and is 3 B-cun (proportional bone cun) the treatment. This scale has been used in a PD study
superior to the prominence of the medial malleolus [35]. [39]. The scale measures a continuous quantitative
variable varying from 0 mm (no anxiety) to 100 mm
(a) Deqi Group. The treatment will be performed after (almost death).
sterilizing the skin on the areas where the needles will be
inserted, with participants lying face up. Using single-use (2) The changes of temperature at SP6 (Sanyinjiao) and
sterile needles (Zhongyan Taihe, Wuxi Jiajian Medical Instru- CV4 (Guanyuan) acupoints (located at the lower
ment Co. Ltd., Jiangsu, China) of 0.30 mm calibre and 40 mm abdomen, 3 B-cun inferior to the centre of the umbili-
length, a needle will be vertically inserted 1 cun in depth and cus, on the anterior median line [35], e.g., in the skin
will be manipulated by lifting-thrusting and twirling methods area corresponding to the uterus) are measured by a
for 30 seconds to achieve Deqi. digital infrared thermographic imaging device (USA,
FLIR Systems Inc., SC640) before and after treatment.
(b) No Deqi Group. Single-use sterile needles (Zhongyan It is demonstrated that acupuncture can affect skin
Taihe, Wuxi Jiajian Medical Instrument Co. Ltd., Jiangsu, temperature at the needling acupoint, and it is feasible
China) of 0.18 mm calibre and 13 mm length will be used. The to detect the changes of the skin temperature by using
treatment will be performed after sterilizing the skin on the a thermographic camera at a certain acupoint during
areas where the needles will be inserted, with participants acupuncture treatment [40].
lying face up. A needle will be vertically inserted 0.1 cun in (3) Adverse events: the possible side effects and adverse
depth. No manipulation will be performed after insertion of reactions during the treatment will be recorded in the
needles to avoid Deqi. case report form (CRF).
Table 1: Data collection at different assessment points. Example of a group of specific subjective sensations which can only be
outcome measurements. felt during acupuncture by both patients and acupuncturists.
Variable T0 T1 It is impossible to randomly allocate participants into a Deqi
or no Deqi group, so there have been no Deqi randomized
VAS-P X X
controlled trials (RCTs). However, whether Deqi can be
VAS-A X X
achieved during acupuncture can be predicted to a certain
ADCAS X extent; therefore, we will randomly divide all participants
Temperature at SP6, CV4 X X into Deqi or no Deqi group. We also employ manipulation
Side effects and adverse reactions X technique to promote Deqi in the Deqi group, while in the
Sociodemographic data X no Deqi group we avoid Deqi according to TCM theory and
Note: VAS-P: visual analogue scale for pain (0–100); VAS-A: visual analogue experts’ experience.
scale for anxiety (0–100); ADCAS: Acupuncture Deqi Clinical Assessment In this study, we will use an important self-designed
Scale; SP6: Sanyinjiao; CV4: Guanyuan; T0: before acupuncture treatment; instrument—Acupuncture Deqi Clinical Assessment Scale
T1: after acupuncture treatment.
(ADCAS). The scale shows the intensity of participants’
sensation, ranging from 0 to 4: 0 (no), 1 (slight), 2 (mild),
2.6. Data Collection. The data required for evaluating the 3 (obvious), and 4 (strong). It will be firstly used as an
influences of Deqi on acupuncture therapy will be collected at instrument for redividing all the included participants in
baseline and the completion of intervention. Data will be reanalysis. Actually, some researchers and their colleagues
obtained via physical measurements. Data collection instru- have attempted to qualify and quantify sensations of Deqi
ments and the study timeline are summarized in Table 1. [41–47] and designed several Deqi scales or questionnaires
A case report form (CRF) has been designed, to include like the Vincent scale, the Park questionnaire, the MGH
the variables of interest, which will be completed by the Acupuncture Sensation Scale (MASS), and Southampton
corresponding researcher at each research center. After the Needling Sensation Questionnaire (SNSQ). Some of the
trial finished, the information obtained will be input to an scales or questionnaires have been used in Deqi trials [48–50]
electronic database, for subsequent statistical analysis. and MASS even has a Chinese edition [51]. However, though
Deqi sensation can be felt by patients and acupuncturists, all
2.7. Data Storage and Confidentiality. All CRFs will be stored the current scales take no consideration of the Deqi sensation
in a locked cabinet at a study office in School of Acupunc- felt by acupuncturists’ hands, which decreases the reliability
ture, Moxibustion and Tuina, Beijing University of Chinese of those scales.
Medicine, and will have a unique identification number. Data Additionally, all the previous scales or questionnaires
will be input to an electronic database, and the access to the derived from a pain questionnaire—McGill Pain Question-
database is restricted to the study team. naire (MPQ) [52], which are not based on TCM theory. Thus,
we designed this new Deqi questionnaire based on the pre-
2.8. Statistical Analysis. All analyses will be performed using vious instruments and especially followed TCM theory and
SPSS 17.0 (SPSS Inc., Chicago, IL). Potential differences rigorous methodology, such as selecting the items included
across study groups on demographic and clinical history in the Deqi questionnaire and testing reliability and validity.
variables will be compared by means of analysis of covari- In fact, the reliability and validity of ADCAS were evaluated
ance (ANCOVA). Repeated-measures analysis of variance by a study with 73 PD patients needled on SP6, in which the
(ANOVA) will be utilized for the analysis of VAS-P and VAS- disease and selected acupoint were the same as in this study,
A scores and temperatures at SP6 (Sanyinjiao) as well as CV4 and it had a good result (the paper will be published later).
(Guanyuan) acupoint. 𝑃 < 0.05 will be denoted as significant. As a subjective sensation, Deqi has no objective measure
Although most participants of the Deqi group will expe- instrument by far. This trial will explore the possibility of
rience Deqi, there will probably be some exceptions that acupoint temperature changing as a sign of Deqi and will be
participants in the group will not experience Deqi. Therefore, devoted to the provision of a tool to determine Deqi.
we will redivide all participants into the real Deqi group and This trial, in accordance with the STRICTA [53] and good
unreal Deqi group according to a self-designed Acupunc- clinical practice guidelines (GCP), will be helpful in explain-
ture Deqi Clinical Assessment Scale (ADCAS) grade after ing the role of Deqi in acupuncture analgesia, provide new
treatment. We will perform a secondary analysis with similar objective evidence for Deqi, and improve the understanding
statistical method to explore whether the real Deqi has of complicated mechanisms of acupuncture in practice.
influences on the effect of acupuncture. We will compare the
first and second analysis results. Conflict of Interests
3. Discussion The authors declare that there is no conflict of interests
regarding the publication of this paper.
The trial is designed to illustrate Deqi effect on acupuncture
treatment. It will estimate Deqi state of solo acupoint and Authors’ Contribution
compare analgesic right after needling. All participants will
be randomly divided into Deqi or no Deqi group. Unlike Yu-qi Liu and Peng Zhang as well as their affiliations con-
other impact factors of acupuncture treatment, Deqi is tributed equally to this work.
Acknowledgments Jianghan University,” Jiang Han Da Xue Xue Bao (Zi Ran Ke Xue
Ban), vol. 38, no. 3, pp. 74–76, 2010.
The authors sincerely thank Professors Lu-fen Zhang and [15] M. I. Ortiz, “Primary dysmenorrhea among Mexican university
Xiao-Xuan Ren at School of Acupuncture, Moxibustion and students: prevalence, impact and treatment,” European Journal
Tuina, Beijing University of Chinese Medicine. The authors of Obstetrics & Gynecology and Reproductive Biology, vol. 152,
also sincerely thank the National Basic Research Program no. 1, pp. 73–77, 2010.
of China (973 Program) (no. 2012CB518506 and no. [16] A. Polat, H. Celik, B. Gurates et al., “Prevalence of primary
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National Natural Science Foundation of China (no. 2009.
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http://dx.doi.org/10.1155/2015/904926
Research Article
Acupuncture for Functional Dyspepsia: A Single Blinded,
Randomized, Controlled Trial
Yulian Jin,1,2 Qing Zhao,2 Kehua Zhou,3 Xianghong Jing,4 Xiaochun Yu,4 Jiliang Fang,2
Zhishun Liu,1 and Bing Zhu4
1
Department of Acupuncture and Moxibustion, Guang An Men Hospital, China Academy of Chinese Medical Sciences,
No. 5 Beixiange Street, Xicheng District, Beijing 100053, China
2
Department of Radiology, Guang An Men Hospital, China Academy of Chinese Medical Sciences, Beijing 100053, China
3
Department of Health Care Studies, Daemen College, Amherst, NY 14226, USA
4
Institute of Acupuncture and Moxibustion, China Academy of Chinese Medical Sciences, Beijing 100700, China
Correspondence should be addressed to Jiliang Fang; fangmgh@163.com and Zhishun Liu; liuzhishun@aliyun.com
Received 11 August 2014; Revised 18 September 2014; Accepted 3 October 2014
Academic Editor: Jian Kong
Copyright © 2015 Yulian Jin et al. This is an open access article distributed under the Creative Commons Attribution License, which
In order to investigate the therapeutic potential of acupuncture on patients with functional dyspepsia (FD), patients were
randomized to receive acupuncture at classic acupoints with manipulations (treatment group) versus acupuncture at nonacupoints
without manipulation (control group) once every other day, three times a week, for one month and were followed up for three
months. The primary outcomes included dyspeptic symptoms, quality of life, and mental status. The secondary outcomes included
the fasting serum gastrin concentration, and frequency and propagation velocity of gastric slow waves. Sixty patients with FD
were included, among whom, four dropped out. After one month’s treatment, patients with FD showed significant improvements
in primary (in both groups) and secondary (in the eight patients of the treatment group) outcomes as compared with baseline
(𝑃 = 0.0078 to <0.0001); treatment group has better outcomes in all primary outcome measures (𝑃 < 0.0001 except for SDS
(𝑃 = 0.0005)). Improvements on dyspeptic symptoms persist during follow-up (better in the treatment group). Acupuncture with
manual manipulation had better effects on improving dyspeptic symptoms, mental status, and quality of life in patients with FD.
These effects may be related to the increased frequency and propagation speed of gastric slow waves and serum gastrin secretion.
1. Introduction visceral sensation, and psychosocial factors have all been

identified as major pathophysiological changes in FD [3, 4].
Functional dyspepsia (FD) is dyspepsia without evidence The prevalence of FD varies between 11% and 29.2% [5]. In
of an organic disease that is likely to explain the cause the United States, FD was found in 29.2% of the population,
[1]. Based on the Rome III criteria, symptoms of FD may and in the United Kingdom the prevalence was 23.8% [6, 7].
include bloating, belching, early satiety, abdominal disten- FD greatly decreases patients’ quality of life as the
sion, nausea, or indigestion during the last three months symptoms, particularly abdominal pain and indigestion,
with symptom onset at least six month ago. These symptoms cause emotional distress, problems with food and drink,
are categorized into epigastric pain syndrome and postpran- and impaired vitality [8]. Patients with FD usually require
dial distress syndrome [2]. Pathophysiological factors which extensive diagnostic procedures and long-term medical care
may cause FD include genetic predispositions, early family which in turn place heavy economic burden on patients
environment, psychosocial factors, abnormal gastric motility, and the society. Management of FD mainly includes lifestyle
visceral hypersensitivity, inflammation, and bacterial flora modification, H. Pylori treatment, acid suppression therapy,
[3]. Particularly, gastrointestinal motor abnormalities, altered prokinetics, antidepressants, and antiflatulents. Despite these
treatment options, treatment for FD often remains unsatis- 2. Material and Methods
factory [9]. The management of FD is challenging especially
when initial drug therapy fails, which is not uncommon [10]. 2.1. Study Design and Setting. This was a single blinded,
Furthermore, besides side effects, traditional drug therapy randomized, controlled trial of manual acupuncture on
has been strikingly shown to have little to no efficacy [11]. classic acupoints versus nonclassic acupoints performed at
For example, benefits from H. Pylori treatment were found the Department of Acupuncture at Guang An Men Hospital,
to be minimal [12]; acid suppression therapy was found to one of the top teaching hospitals for TCM education in
be suboptimal with no apparent effects on dysmotility-like China. Hospital ethics committee approved the study proto-
dyspepsia [13]. col. Participants were recruited through advertisements on
From a traditional Chinese medicine (TCM) perspec- local newspapers, posters, and hospital website and signed
tive, FD is characterized by disrupted qi flow inside the informed consent before study participation.
middle energizer due to external pathogenic factors [14]. An investigator who was not involved in acupuncture
The middle energizer refers to spleen and stomach which procedures and data analyses was responsible for the genera-
are responsible for food transformation and transportation. tion of a random number table, based on which, participants
Kidney is responsible for bone health and the generation were allocated to receive either classic acupoint (treatment
of bone marrow; excessive physical work consumes kidney group) or nonclassic acupoint (control group) acupuncture
energy. Meanwhile, excessive mental work consumes blood treatments. Participants were blinded to acupuncture proce-
and causes imbalance of emotion. Blood is controlled by dures.
heart and emotion is regulated by liver. Thus, treatment in
TCM including acupuncture should aim to facilitate qi and 2.2. Participants. For inclusion, patients have to fulfill the
blood circulation in meridians related to these organs and Rome III diagnostic criteria for FD. For the last three months
thus normalizes patient’s status of health. With the guidance with symptom onset at least six months ago, patient has
of these diagnostic and therapeutic principles, therapeutic one or more of the following: (1) bothersome postprandial
effectiveness of acupuncture for abdominal pain, abdominal fullness; (2) early satiation; (3) epigastric pain; (4) epigastric
distension, bloating, nausea, and others was well documented burning; and (5) no evidence of structural disease (including
in various TCM classics and has been reported in research at upper endoscopy) that is likely to explain the symptoms.
studies [15–20]. In addition, the following criteria were also met: no mental
Besides normalization of qi and blood in the affected disorders; would otherwise be healthy; age 18 to 70; non-
meridians, modern understandings of these results also lie pregnant; and one week prior to and during participation,
in pathophysiological research studies, in which researchers cessation of all medication related to the gastrointestinal
found that acupuncture in patients with FD could accelerate system, which may include but not be limited to gastric
solid gastric emptying [17], increase plasma level of neu- suppression drugs, prokinetics, H. Pylori eradication agents,
ropeptide Y but not motilin [18], and induce deactivation and antidepressants.
of the brainstem, anterior cingulate cortex (ACC), insula, Eight patients in the treatment group also signed in
thalamus, and hypothalamus in the human body [21]. In for assessment of gastrin concentration and frequency and
addition, acupuncture was also found to enhance normal propagation velocity of gastric slow waves. To measure the
gastric myoelectrical regularity in both healthy people and differences between patients with FD and healthy adults
patients with diabetic gastric dysrhythmia [22, 23], alters the in these assessments, eight healthy volunteers were also
frequency of gastric slow waves in healthy volunteers [24], included to match with these eight patients with FD in the
and accelerates solid gastric emptying in diabetic gastropare- present study.
sis [25].
Acupuncture seems to be a promising treatment for FD;
however, the aforementioned clinical trials did not investigate 2.3. Treatment Protocol. All patients included were random-
the effects of acupuncture on emotional symptoms [15– ized into two groups: classic acupoint (treatment) or nonacu-
25], the prevalence of which has been found to be high point (control) groups. For the treatment group, acupoints
in patients with FD [26]; placebo effect which is common ST36 and KI3 were used in every group members; additional
in both patients with FD and acupuncture procedures will acupoints of GB4, PC6, and HT7 may also be used based on
likely add more uncertainties in the therapeutic effectiveness pattern recognition of symptoms. Based on TCM theories,
of acupuncture [27, 28], and, finally, not all of the studies ST36 was used to invigorate functions of the stomach and
performed acupuncture procedures based on TCM principles spleen; KI3, the Yuan-source acupoint, was used to invigorate
including the meridian theories, such as an emphasis on Deqi functions of the kidney. ST36 and KI3 function together to
sensations. restore the normal qi flow inside the stomach and spleen
In the present study, we aimed to determine (i) the effect meridian. In addition, for patients with obvious depression,
of acupuncture on dyspeptic symptoms, quality of life, and anxiety, or insomnia symptoms, GB41 was used to restore
mental status in patients with FD; (ii) the effect difference liver function, and PC6 and HT7 were used to nourish the
between classic acupuncture based on TCM principles and heart to resume balance of the mind. Classic acupoints were
acupuncture on nonacupoints; and (iii) effects of classic localized according to the 2008 World Health Organization
acupuncture on serum gastrin concentration and frequency standards [29]. For the treatment group, needle insertion
and propagation velocity of gastric slow waves. was perpendicular with a depth of about 25 mm. In order to
reach an optimal response which is defined as Deqi sensa- Quality of life was measured by the short-form 36
tions including soreness, heaviness, fullness, propagation of (SF-36) questionnaires [33]. Mental statuses of patients were
needling sensation, and/or adjacent muscle twitching [30], evaluated via Zung Self-Rating Depression Scale (SDS) [34]
moderate combined acupuncture manipulation of lifting, and Self-Rating Anxiety Scale (SAS) [35]. Scoring of these
thrusting, and twirling with a frequency of 60–120 times/min standardized assessments followed guidelines published in
was performed. These acupuncture manipulation techniques the Manual of Standardized Assessment Tools in Behavioral
were performed continually to reach one to three times of Medicine [36]. SF-36 measures Quality of Life (QoL) across
Deqi sensation (with a short interval between Deqi sensations eight domains; score of each domain = [(actual raw score −
if more than once during the first two minutes); then, the lowest possible raw score)/raw score range] × 100. For the
needle was removed. If no Deqi sensation was obtained SDS score, the following equation was used: SDS Index =
during the first two minutes, acupuncture needle was then left Raw Score × 1.25. Grading of SDS is as the following: SDS
in place for 20 to 60 minutes, and one acupuncture manipu- Index less than 53 points is considered normal, 53 to 62 as
lation was applied right before needle removal regardless of mild depression, 63 to 72 as moderate depression, and 73 and
Deqi sensation. higher as severe depression [36]. For the grading of SAS, the
For the control group, nonclassic acupoints in different following categories were used: normal range (less than 50),
dermatomes but close proximity of the aforementioned mild anxiety (50 to 59), moderate anxiety (60 to 69), and
acupoints were used in the distal portion of extremities severe anxiety (70 and higher) [36].
correspondingly. KI3, ST36, and GB41 are located in the The secondary outcomes include fasting serum gastrin
L4, 5, and S1 dermatome; thus nonclassic acupoints located concentration and frequency and propagation velocity of
inside anterior thigh (L2 and L3 dermatome) were used. PC6 gastric slow waves. These measurements were performed in
and HT7 are located inside the C7, 8, and T1 dermatome; the eight patients with FD in the treatment group before
thus nonclassic acupoint in the anterior antebrachium (C5 and after treatment, but only once in healthy volunteers. A
dermatome) was used. In the control group, needle insertion fasting venous blood sample was drawn from the basilic vein
was perpendicular with a depth of two to three millimeters prior to breakfast early in the morning. About three milliliters
with needle retention of 20 minutes but no acupuncture of the blood sample was sent to Peking Union Medical
manipulations. College Hospital for measurement of serum gastrin levels.
Treatments in both groups were implemented once every Meanwhile, the participant was given 120 mL 80% (w/v)
other day, three to four times a week for one month. barium sulfate suspension (Qingdao Dongfeng Chemical Co.
All patients were then followed up for three months. Ltd., Shandong, China). Participants were then placed in
All acupuncture procedures were performed by the same a supine position. Using Prestige digital X ray (GE, USA),
acupuncturist who had more than six years’ clinical experi- gastric mucosa was observed; then, a Chinese coin of fifty
ences. Huatuo brand needles (Φ 0.35 mm × 25 mm, manu- cents was placed on top of the skin over the stomach of the
factured by Suzhou Medical Appliance manufactory, Jiangsu, participant, and gastric motions around the gastric antrum
China) were used for all acupuncture procedures. were video recorded for one minute while the participant
was in a standing position. Frequency of gastric slow waves
was directly counted as the number of waves that passed
2.4. Outcome Assessment. The primary outcomes of the study through the gastric antrum in one minute. Propagation
included dyspeptic symptoms, quality of life, and mental velocity of gastric slow waves was assessed by the time interval
status. For dyspeptic symptoms, we used the four cardinal between two consecutive waves that passed through the
dyspeptic symptoms and their corresponding assessments as gastric antrum.
reported in the Chinese version Nepean Dyspepsia Index Safety evaluation includes possible hematoma, local
(NDI) [31, 32]. The intensity, frequency, and level of inter- infection, fainting, and severe pain during and after acupunc-
ference of postprandial fullness, early satiety, epigastric pain, ture. In addition, other conditions which warrant cessation of
and epigastric burning sensation were rated. Intensity of acupuncture treatment or withdrawal from the study if any
each symptom was graded and scored as the following: 0, were also documented and analyzed.
absent; 1, mild; 2, moderate; 3, severe; 4, critical. Frequency
of each symptom was also graded as follows: 0, absent; 1,
occasionally (1-2 days/week); 2, sometimes (3–5 days/week); 2.5. Statistical Analysis. The statistical analysis was per-
3, frequently (every day, but intermittent symptoms), 4, formed by two independent statisticians. Results were com-
continuous symptoms. Level of interference of each symptom pared between the two statisticians. Differences, if any, were
was scored and graded as the following: 0, none; 1, mild discussed and the statistic test was reperformed until a
interference; 2, moderate interference; 3, severe interfer- consensus was reached between the two statisticians. The
ence; 4, critic interference. The number in front of each statisticians were blinded to treatments and study protocol.
grading indicates the score of the corresponding symptom; All results including baseline characteristics were based on
the score for each symptom in the checklist of cardinal per-protocol (PP) analyses. Statistical Analysis System (SAS),
dyspeptic symptoms was calculated by adding its scores in the version 6.12, was used and a significance level was set at 𝑃 <
corresponding frequency, severity, and level of discomfort; 0.05.
dyspeptic symptom sum score (DSSS) is the sum score of the For comparisons of baseline values, chi square test was
four symptoms in the checklist. used to explore gender differences; 𝑡-test was used to explore
Assessed for eligibility

(n = 88)
Excluded (n = 28)
∙ Peptic ulcer (n = 4)
∙ Superficial gastritis (n = 7)
∙ Atrophic gastritis (n = 6)
∙ Gastroesophageal reflux disease (n = 3)
∙ Cholecystitis (n = 2)
∙ Hashimoto thyroiditis (n = 1)
∙ Diabetes mellitus (n = 2)
∙ Severe coronary artery disease (n = 2)
∙ Older than 70 (n = 1)
Patients who accepted

acupuncture
treatment (n = 60)
Treatment group (n = 30) Control group (n = 30)

Dropout (n = 2) Dropout (n = 2)
Figure 1: Flow chart of study participation.
differences in the duration of the disease; Wilcoxon rank sum treatment was noneffective and withdrew from participation
test was used in all comparisons of primary and secondary (Figure 1).
outcome measures. All quantitative data including subjective The treatment group consists of 11 males and 17 females
scores were expressed with mean ± SD. with an age range between 23 and 65 years old and disease
history of one to 40 years. The control group consists of 10
males and 18 females with an age range between 24 and 66
3. Results years old and disease history of one to 40 years. Prior to
From July, 2010, to January, 2011, a total of 88 patients with participation, no significant differences were found between
dyspeptic symptoms visited the Department of Acupunc- these two groups in terms of gender, age, length of disease
ture at Guang An Men Hospital in Beijing. Twenty-eight history, dyspeptic symptom sum scores, and SF-36 score
patients were excluded from the present study due to the (Table 1).
following reasons: peptic ulcer (four patients), superficial
gastritis (seven patients), atrophic gastritis (six patients), 3.1. Primary Outcomes. At baseline, the prevalence of the four
gastroesophageal reflux disease (three patients), cholecystitis symptoms of postprandial fullness, early satiety, epigastric
(two patients), Hashimoto thyroiditis (one patient), diabetes pain, and epigastric burning sensation in these 56 patients
mellitus (two patients), severe coronary artery disease (two were 98.2%, 71.4%, 76.8%, and 58.9%, respectively; the scores
patients), and older than 70 (one patient). Sixty patients for each symptom were six to nine points with a severity of
were included and randomly assigned to either the treatment disease rated moderate to severe.
group or the control group. Of these 60 patients, 56 patients After one month’s treatment, as compared with baseline
completed the study and four patients (two from each group) values, significant differences were found in both treatment
dropped out from the study (dropout rate: 6.67%) after and control groups in the dyspeptic symptom sum score, the
the second visit. In the treatment group, one patient could scores of postprandial fullness, early satiety and epigastric
not tolerate the acupuncture Deqi sensations upon needle pain, SDS score, and SF-36 score. Additionally, as compared
manipulation, and the other patient in the treatment group with baseline, significant differences were also found in the
had transportation difficulties. In the control group, the score of epigastric burning sensation and SAS score of the
two patients directly stated to the therapist saying that the treatment group but not the control group. 𝑃 values were
Table 1: General characteristics of patients with FD prior to participation.
Gender
Groups Cases (𝑛) Age (year) Disease duration (years) Dyspeptic symptom sum score SF-36 score
Male (𝑛) Female (𝑛)
Treatment 28 11 17 49.29 ± 10.32 12.20 ± 12.20 24.32 ± 8.28 52.51 ± 13.94
Control 28 10 18 48.25 ± 11.40 12.11 ± 10.20 24.79 ± 7.48 54.06 ± 16.41
𝑃 value 0.7825 0.7229 0.6145 0.8265 0.7043
<0.0001 for all the significant intragroup comparisons except The results of the present study add further credence to the
epigastric pain, SF-36, and SDS in the control group, for use of these acupoints.
which 𝑃 values were 0.0078, 0.0099, and 0.0002, respectively.
As compared with the control group, treatment group has
better outcomes in all primary outcome measures. 𝑃 values 4.2. Acupuncture Manipulations. Acupuncture Deqi serves
for these intergroup comparisons were all <0.0001 except for as the foundation or premise for the therapeutic effects of
SDS (𝑃 = 0.0005) (Table 2). acupuncture treatment [30]. Although theoretical research
At three months’ follow-up, DSSS was recalculated for all articles highlight the importance and the components of
participants. As compared with baseline values, significant Deqi, not many researchers emphasized Deqi in their reports
differences were found in both groups in terms of DSSS (𝑃 < of acupuncture clinical trials. The reason for lacking of
0.0001). Meanwhile, the treatment group, as compared with information regarding Deqi may be due to the following
the control group, had better long-term outcomes in terms of reasons: the authors of the reports did not document it and
DSSS (𝑃 < 0.0001) (Table 3). the clinicians did not pay extra attention to the importance of
Deqi during the studies. In addition, as electroacupuncture
3.2. Secondary Outcomes. Values of preprandial serum gas- becomes more and more popular, the evaluation of Deqi is
trin concentration and frequency and propagation velocity more difficult due to the mixture of electric therapy sensa-
of gastric slow waves in healthy volunteers and patients with tion with sensations from acupuncture itself. Nonetheless,
FD were provided in Table 4. As compared with healthy report of Deqi in clinical trials reflects more the standard
volunteers, patients with FD had lower serum gastrin concen- acupuncture treatment in clinical practice. In the present
tration and less frequent and slower propagation velocity of study, manual acupuncture manipulation was stopped and
gastric slow waves (𝑃 = 0.0081, 0.0008, 0.0279, resp.) at base- the needle was removed upon Deqi arrivals in the treatment
line. After one month’s treatment, patients with FD showed group. This acupuncture treatment protocol guarantees not
significant improvement in serum gastrin concentration and only the Deqi sensations thus the clinical efficacy but also
frequency and propagation velocity of gastric slow waves safety of acupuncture treatments. The results of the present
(𝑃 = 0.0002, 0.0078, and 0.0180, resp.), and no significant study indicate that traditional acupuncture with the emphasis
difference was found in these secondary outcome measures of Deqi manipulations has better therapeutic results than
between healthy volunteers and patients with FD (Table 5). acupuncture on nonacupoints without Deqi manipulations.
3.3. Side Effects. No serious side effects occurred. One patient 4.3. Outcome Measurement. As psychosocial factor is a com-
in the treatment group withdrew from the study secondarily mon cause of FD and many patients of FD have anxiety
to intolerance to the needling sensations upon acupuncture or depression issues, measurements of these psychological
manipulation. symptoms are of great importance in the evaluation of clinical
management of FD [1, 26]. Zung Self-Rating Depression
4. Discussion Scale (SDS) and Self-Rating Anxiety Scale (SAS) have a high
reliability and validity in assessing psychological symptoms
4.1. Selection of Acupoints. The use of classic acupoint of in patients [34, 35]. The improvement of SDS score and SAS
ST36 in the present study is well-supported by former score in the treatment group of the present study indicates
research studies [15–25], so was the use of PC6 [15–19, 24]. that acupuncture has positive impacts on the psychological
In previous research studies, researchers mainly considered aspects of patients with FD. Psychological effects of acupunc-
the pathophysiological relationship between the meridians ture may be caused by placebo effects [28, 37]; however,
or organs of liver and spleen, heart and spleen, or spleen as acupuncture also demonstrated therapeutic effects on
and kidney; acupoint of the kidney meridian is barely used psychological diseases [38, 39], we should increase our trust
for FD in these research studies [15–25]. In the present on the positive benefits of acupuncture on psychological
study, we used KI3 based on the analysis of all the patho- symptoms of patients. To our best knowledge, no studies
physiological relationships between and among organs and have explored the effects of acupuncture on psychological
meridians related to FD symptoms. These diagnostic and symptoms of FD. Thus, the present study will facilitate our
therapeutic principles would be a more realistic reflection understanding of the therapeutic effectiveness of acupunc-
of individualized acupuncture treatment in clinical practice. ture in FD.
Table 2: Scores of dyspeptic symptoms, quality of life, and mental status before and after the treatment.
Items Groups 𝑁 Baseline After treatment Difference Improvement rate 𝑃 value

Treatment 28 9.00 ± 2.09 1.57 ± 2.28 7.43 ± 2.47 82.56% <0.0001
PF Control 27 8.89 ± 2.39 6.22 ± 2.59 2.67 ± 1.88 30.03% <0.0001
IGC <0.0001
Treatment 19 9.74 ± 1.91 0.42 ± 1.43 9.32 ± 1.97 95.69% <0.0001
ES Control 21 8.43 ± 2.87 6.05 ± 2.52 2.38 ± 1.80 28.23% <0.0001
IGC <0.0001
Treatment 21 6.81 ± 2.23 0.48 ± 1.03 6.33 ± 2.31 92.95% <0.0001
EP Control 22 7.41 ± 3.02 6.32 ± 3.41 1.09 ± 1.82 14.71% 0.0078
IGC <0.0001
Treatment 16 6.31 ± 2.39 0.50 ± 1.55 5.81 ± 2.17 92.08% <0.0001
EBS Control 17 6.71 ± 2.78 6.47 ± 3.00 0.24 ± 0.56 3.58% 0.25
IGC <0.0001
Treatment 28 24.32 ± 8.28 2.50 ± 3.28 21.80 ± 8.24 89.72% <0.0001
DSSS Control 28 24.79 ± 7.48 19.40 ± 8.23 5.36 ± 3.29 21.62% <0.0001
IGC <0.0001
Treatment 28 52.50 ± 13.94 70.00 ± 12.54 17.00 ± 14.04 33.52% <0.0001
SF-36 Control 28 54.00 ± 16.41 56.00 ± 13.42 2.88 ± 8.74 5.33% 0.0099
IGC <0.0001
Treatment 28 57.96 ± 9.55 45.60 ± 8.75 12.30 ± 9.89 21.33% <0.0001
SDS Control 28 57.60 ± 11.84 54.00 ± 10.80 3.50 ± 5.92 6.07% 0.0002
IGC 0.0005
Treatment 28 52.30 ± 10.48 42.30 ± 6.22 10.00 ± 10.22 19.11% <0.0001
SAS Control 28 52.36 ± 9.67 52.20 ± 7.98 0.11 ± 4.89 0.21% 0.8533
IGC <0.0001
PF: postprandial discomfort; ES: early satiety; EP: epigastric pain; EBS: epigastric burning sensation; DSSS: dyspeptic symptom sum score; SF-36: short-form
36 questionnaire; SDS: Self-Rating Depression Scale; SAS: Self-Rating Anxiety Scale; IGC: intergroup comparison.
Table 3: Dyspeptic symptom sum score at baseline and during follow-up.
IND Groups 𝑁 Baseline Follow-up Difference Improvement 𝑃 value

Treatment 28 24.32 ± 8.28 1.68 ± 2.36 22.60 ± 8.68 93.09% <0.0001
DSSS Control 28 24.79 ± 7.48 16.43 ± 7.41 8.36 ± 6.58 33.92% <0.0001
IGC <0.0001
DSSS: dyspeptic symptom sum score; IGC: intergroup comparison.
Quality of life is a heavy emphasis of the clinical manage- reported by Park et al. [19]. Significant superiority of classic
ment of all kinds of disorders. In the present study, the use acupoint acupuncture to nonclassic acupoint acupuncture
of modified NDI is well-supported by its high reliability and was found in both studies by Ma et al. [20] and Zeng et al.
validity in patients with dyspeptic symptoms [31, 32]. NDI [21]; however, they did not report changes of subcategories of
measures dyspepsia symptoms and dyspepsia-specific health- NDI in both groups which makes the analysis difficult. Park
related QOL (H-QOL). Outcome measurements utilizing et al. [19] did not find difference between classic acupoint
NDI, SAS, and SDS will likely better capture the charac- acupuncture and nonclassic acupoint acupuncture except for
teristics of acupuncture effects on FD. The improvement of pressure and cramps in upper abdomen (better results in the
NDI in the present study concurs with results from other classic acupoint acupuncture group). The differences may be
acupuncture researchers regarding acupuncture treatment due to control group treatment. In the study by Park et al.
for FD [15–21]. Interestingly, the control group in which [19], dermatome information between classic acupoint and
acupuncture was used in nonclassic acupoints also induced nonclassic acupoint was not included in consideration upon
significant changes in dyspeptic symptoms except for epigas- the design of control group.
tric burning sensations and SAS score. These results partially FD, like other diseases, is characterized by its objective
concur with the results reported by Ma et al. [20] and physiological changes and subjective symptoms; thus, a
Zeng et al. [21]; however, the results differ from the results thorough evaluation of FD should simultaneously include
Table 4: Serum gastrin concentration and frequency and propagation velocity of gastric slow waves in patients with functional dyspepsia
and healthy adults.
Items Baseline (𝑛 = 8) After treatment (𝑛 = 8) Healthy adults (𝑛 = 10)

Gastrin (pg/mL) 25.93 ± 5.90 44.40 ± 6.26 47.65 ± 20.21
FGSW (n/min) 2.49 ± 0.64 3.11 ± 0.14 3.11 ± 0.13
PVGSW (s) 24.25 ± 4.95 19.75 ± 2.05 19.41 ± 0.93
FGSW: frequency of gastric slow waves; PVGSW: propagation velocity of gastric slow waves.
Note: propagation velocity of gastric slow waves was assessed by the time interval between two consecutive waves that passed through the gastric antrum.
Table 5: Comparisons of serum gastrin concentration and frequency and propagation velocity of gastric slow waves before and after treatment
as well as between patients with functional dyspepsia and healthy adults.
Items Baseline versus healthy Baseline versus after After treatment versus
adults treatment healthy adults
Gastrin 0.0081 0.0002 0.6401
FGSW 0.0008 0.0078 1.0000
PVGSW 0.0279 0.0180 0.6713
FGSW: frequency of gastric slow waves; PVGSW: propagation velocity of gastric slow waves.
Note: propagation velocity of gastric slow waves was assessed by the time interval between two consecutive waves that passed through the gastric antrum.
“Baseline versus after treatment” refers to patients with functional dyspepsia only.
these two aspects. Changes in gastric motility, mainly gastric An accepted mechanism of acupuncture on the func-
hypomotility or dysrhythmias, play an important role in the tions of the gastric system is related to its effects on
pathophysiology of FD [1–4, 13]; thus, an objective mea- the autonomic nervous systems, which Takahashi [15]
surement of gastric motilities is of great importance in the summarized as follows: acupuncture at the lower limbs
evaluation of FD. Barium sulfate radiography of the GI system (ST36) causes gastric muscle contraction via stimulating
provides a direct visual observation of the frequency and the somatoparasympathetic pathway whereas acupuncture
propagation velocity of gastric slow waves and thus objective at the upper abdomen causes gastric muscle relaxation via
measurements of acupuncture effects on FD. Gastrin is a stimulating the somatosympathetic pathway. As both main
peptide hormone that stimulates the secretion of gastric acid acupoints KI3 and ST36 used in the present study are
by the parietal cells of the stomach and aids in gastric motility. located in the lower extremities, the result of enhanced
Results from recent research studies indicate that abnormal gastric motility is likely to be caused by activation of the
gastrin level is a possible contribution factor of FD with somatoparasympathetic pathway increasing the secretion of
gastric dysmotility [4, 40, 41]. The results of gastrin level gastrin and other hormones.
in the present study are consistent with the hypothesis as Furthermore, acupuncture has also been found to induce
patients with FD show decreased preprandial gastrin level changes in cerebral cortex activities of patients with FD [21].
[4, 41]. However, He et al. [40] did not find any difference in Consequently, we hypothesize that effect of acupuncture on
preprandial but postprandial gastrin levels (higher in patients the gastrointestinal system is related to its effects on the
with FD) between patients with FD and healthy volunteers. peripheral nervous system, central nervous system, and the
In the present study, observation of less frequent and slower endocrine systems related to the GI tract. However, to prove
propagation velocity of gastric slow waves in patients with the specific causal relationship among these systems, further
FD via barium sulfate radiography also indicates a decreased research studies are needed.
level of gastrin. Nonetheless, changes of gastrin and gastric
motility in patients with FD deserve further research. In
the present study, classic acupuncture was found to increase 5. Limitations
preprandial gastrin level and enhance gastric motility of
As blinding is difficult in acupuncture studies, the establish-
patients with FD to reach similar levels as healthy volunteers.
ment of a blank control group seems impossible. Although
These results are consistent with our findings in improvement
nonclassic acupoint acupuncture procedures were used as
of dyspeptic symptoms.
control in the present study, they are still acupuncture
procedures; thus we could not rule out the cofounding
4.4. Therapeutic Mechanism of Acupuncture. Former re- factor of needling and placebo effects in the present study.
search studies in human beings indicate that acupuncture This study is performed at one clinical center with one
could accelerate solid gastric emptying [18, 25] and enhance acupuncturist on a relatively small sample; the results of
percentage of normal gastric slow waves [22, 23]. The present the present study may not well characterize the response of
study showed similar results in increasing gastric motility patients with FD to acupuncture treatments. In addition, the
as demonstrated by increased frequency and propagation analysis of the results did not include patients who dropped
velocity of gastric slow waves in patients with FD. out; data processing based on per protocol population may
decrease the credence of the results. To better capture the Reviews Gastroenterology & Hepatology, vol. 10, no. 3, pp. 187–
response of patients with FD to acupuncture, further large 194, 2013.
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http://dx.doi.org/10.1155/2015/310591
Research Article
Discovery of Acupoints and Combinations with Potential to
Treat Vascular Dementia: A Data Mining Analysis
Shuwei Feng,1 Yulan Ren,1 Shilin Fan,2 Minyu Wang,1 Tianxiao Sun,1
Fang Zeng,1 Ping Li,2 and Fanrong Liang1
1
Chengdu University of Traditional Chinese Medicine, Chengdu, Sichuan 610075, China
2
Southwest Petroleum University, Chengdu, Sichuan 610500, China
Correspondence should be addressed to Ping Li; dping.li@gmail.com and Fanrong Liang; acuresearch@126.com
Received 18 December 2014; Revised 16 February 2015; Accepted 30 April 2015
Academic Editor: Zhixiu Lin
Copyright © 2015 Shuwei Feng et al. This is an open access article distributed under the Creative Commons Attribution License,
The prevalence of vascular dementia (VaD) is high among the elderly. Acupuncture, a popular therapeutic method in China,
can improve memory, orientation, calculation, and self-managing ability in VaD patients. However, in clinical acupuncture and
acupuncture research, the selection of acupoints to treat VaD remains challenging. This study aimed to discover acupoints and
acupoint combinations with potential for VaD based on data mining. After database searching and screening for articles on clinical
trials evaluating the effects of acupuncture on VaD, 238 acupuncture prescriptions were included for further analysis. Baihui (GV
20), Sishencong (EX-HN 1), Fengchi (GB 20), Shuigou (GV 26), and Shenting (GV 24) appeared most frequently in the modern
literature and are potential acupoints for VaD. Combinations between Baihui (GV 20), Sishencong (EX-HN 1), Fengchi (GB 20),
Shenting (GV 24), Shuigou (GV 26), and Zusanli (ST 36) were most frequent and represent potential combinations for VaD
treatment. These results provide a reference for the selection and combination of acupoints to treat VaD in clinical acupuncture
and acupuncture research.
1. Introduction Vascular dementia (VaD) refers to cognitive impairment

caused by changes in the blood circulation of the brain [5].
The selection of acupoints plays a critical role in the therapeu-
Its clinical manifestations include confusion or short-term
tic effects of acupuncture. However, the selection of proper
memory problems, wandering, getting lost in familiar places,
acupoints remains challenging, contributing to the limited
walking with rapid and shuffling steps, losing bladder or
therapeutic effects and application of acupuncture. Data
mining has been used to discover potential acupoints from bowel control, laughing or crying inappropriately, difficulty
the expansive relevant literature. This method has been used in following instructions, and problem with counting money
to suggest acupoints on the Shaoyang Meridian for migraine and conducting monetary transactions. At the late stage, VaD
treatment based on their high frequency in the literature [1]. patients may have severe impairment of basic activities of
Based on the results of data mining, a subsequent clinical trial daily living and lack the capacity to make appropriate deci-
confirmed that acupuncture was effective for the treatment sions regarding their choices and preferences [6]. A recent
of migraine and that acupoints on the Shaoyang Meridian population-based survey reported that the prevalence of VaD
were more effective than acupoints on other meridians [2]. among individuals aged 65 years and older was 1.5% [7]. It
Data mining has also been used to discover potential Chinese has been predicted that dementia will affect 80 million people
herbs for the effective treatment of specific diseases [3, 4]. worldwide by 2040 [8]. The annual cost of care per patient
These results support data mining as a promising method to is estimated to be US$17,000–55,200 for severe dementia,
discover acupoints with potential for treating diseases. placing a heavy economic burden on families and society [9].
Acupuncture, a primary therapeutic method in tradi-

tional Chinese medicine (TCM), can improve memory, 2121 records identified through database searching
orientation, calculation, and self-managing ability in VaD (CNKI: 892, CBM: 1084, PubMed: 145)
patients [10–12]. The therapeutic effects of acupuncture
are achieved via multiple pathways, including antioxidative
effects, antiapoptotic effects, and neurotrophic effects [11,
13–15]. However, acupoint selection remains a challenge in
the use of acupuncture to treat VaD. According to our
preliminary statistics, more than 100 acupoints distributed
in 13 meridians have been recorded in the modern literature 238 acupuncture prescriptions included in data
for the treatment of VaD. The most effective acupoints for
mining and network analysis after screening
the treatment of VaD and the selection of acupoints for
combination remain to be elucidated, representing a major
limitation for clinical therapeutic effects and the application
of acupuncture for VaD.
To shed some light on the selection of acupoints and
acupoint combinations to treat VaD in clinical acupuncture
and acupuncture research, this study aimed to discover Data preprocessing: extraction of information about
acupoints and acupoint combinations that have potential to
titles, journals, interventions, and main acupoints
treat VaD via data mining.
and standardization of acupoint names
2. Materials and Methods
The flow of information through the various phases of data
mining is illustrated in Figure 1.
2.1. Inclusion Criteria for Considering Acupoint

Data processing: analysis of frequencies of
Prescriptions for Data Mining
acupoints and meridians, confidence of acupoint
2.1.1. Types of Studies. Clinical trials evaluating the effect of
TCM acupuncture with or without randomization methods combination, and network properties
were included. Trials with or without controls were also
included. The control interventions included no treatment,
Figure 1: Flow of information through the different phases of data
sham acupuncture, Western medicine, TCM herbs, non- mining.
traditional acupuncture, and TCM acupuncture containing
another acupoint prescription which is different from the one
in observation group. Language was restricted to Chinese and
English. Western medicine, TCM herbs, nontraditional acupuncture,
and TCM acupuncture containing another acupoint pre-
2.1.2. Types of Participants. Clinical trials involving adult scription which is different from the one in observation
participants diagnosed with VaD were included. group. If the studies compared the therapeutic effects of
different acupoint prescriptions, the most effective acupoint
2.1.3. Types of Interventions. Clinical trials evaluating TCM prescription was included.
acupuncture were included. Acupuncture can be used
alone or in combination with other types of interventions. 2.2. Exclusion Criteria
TCM acupuncture involves inserting needles into traditional
meridian acupoints and extraordinary acupoints. Electrical 2.2.1. Types of Studies. Case reports, reviews, systematic
stimulation of the needles may be used. Trials using moxi- reviews, and meta-analyses were excluded.
bustion alone or as a cointervention with acupuncture were
also included. 2.2.2. Types of Participants. Trials evaluating the therapeutic
effect of acupuncture for Alzheimer’s disease, traumatic
2.1.4. Effectiveness of Acupoint Prescriptions. Acupoint pre- dementia, and other subtypes of dementia were excluded.
scriptions for the disease and not particular syndromes Studies on animals were also excluded.
of VaD were included. There should be statistical differ-
ences in symptoms between before and after acupuncture. 2.2.3. Types of Interventions. Trials stimulating Ashi points
In a controlled trial, patients treated with acupuncture alone were excluded. Trials of dry needling or trigger point
alone or in combination should receive greater benefit than therapy, therapies that are based on principles of western
patients who do not receive acupuncture therapy. The control anatomy and physiology, were excluded. Trials of laser
interventions included no treatment, sham acupuncture, acupuncture and noninvasive electrostimulation (e.g., using
electrodes on the skin rather than needles or moxibustion to and support of acupoint combinations. According to the
stimulate the acupoints) were excluded to limit the focus to definition of association rules mining [18], the following
TCM acupuncture. Trials evaluating acupressure, a form of can be a statement of association rules mining for acupoint
massage, were excluded as well. Finally, trials of micropunc- combination. Let 𝐼 = {𝑖1 , 𝑖2 , . . . , 𝑖𝑚 } be a set of acupoints.
ture were excluded because micropuncture is a nontraditional Let 𝐷 be a set of acupoint prescriptions, where each acupoint
acupuncture practice that is based on the principle that the prescription 𝑇 is a set of acupoints such that 𝑇 ⊆ 𝐼. Associated
head (or ear, nose, eye, abdomen, ankle, etc.) is a microsystem with each acupoint prescription is a unique identifier, called
of the entire body. TID. An acupoint prescription 𝑇 contains 𝑋, a set of some
acupoints in 𝐼, if 𝑋 ⊆ 𝑇. The rule 𝑋-𝑌 has support 𝑠 in the
2.2.4. Effectiveness of Acupoint Prescriptions. Acupoint pre- acupoint prescription set 𝐷 if 𝑠% of acupoint prescriptions in
scriptions for a particular syndrome of VaD were excluded. 𝐷 contain 𝑋 ∪ 𝑌.
Acupoint prescriptions with no statistical improvement of
symptoms were also excluded. When the therapeutic effects 2.6.3. Measurement of Network Properties. Community struc-
of different acupoint prescriptions were compared in a study, ture is a common characteristic of complex networks and
all acupoint prescriptions except the most effective one were is characterized by more dense internal connections within
excluded. groups of nodes than with the rest of the network. In this
study, a hierarchical agglomeration was adopted to detect
2.3. Searching Methods for Identification of Studies community structure according to Clauset et al. [19]. We also
focused on investigating the set of the most influential nodes
2.3.1. Data Sources. PubMed (http://www.pubmed.com (1966 in acupoint networks of VaD, defined as the nodes with the
to 2012)), the Chinese BioMedicine Database (CBM) (http://
highest 𝑘-core value [20]. The 𝑘-core method is predomi-
www.sinomed.ac.cn (1978 to 2012)), and China National
nantly used in analyzing social networks. We employed the
Knowledge Infrastructure (CNKI) (http://www.cnki.net (1912
𝑘-core method to obtain the cores of different acupoints. The
to 2012)) were searched for modern literature on acupuncture
𝑘-core method was implemented as follows. First, all 1-degree
treatment for VaD.
nodes were removed, and the nodes were further pruned until
2.3.2. Searching Strategy. The searching strategy used the no 1-degree nodes remained. The remaining nodes formed
following key words: (I) “acupuncture” OR “electroacupunc- the 2-core node set. The pruning process was repeated in a
ture” OR “moxibustion” OR “meridian” OR “acupoint”; (II) similar manner for other nodes in the network assigned to
“dementia” OR “vascular dementia” OR “Alzheimer’s disease.” the corresponding cores (denoted by 𝑘s). The nodes with the
The searching strategy included literature on acupuncture largest 𝑘-core value were defined as the network core. The
treatment for Alzheimer’s disease (AD) because the modern degree of each acupoint was also analyzed to measure the
literature on acupuncture for VaD overlaps with that on AD. involvement of the node in the network. The degree refers to
the number of nodes to which a focal node is connected [21].
2.4. Data Collection. Two reviewers independently screened Betweenness centrality was also used to analyze an acupoint’s
the title and abstract of every record retrieved from the centrality in the network. Centrality is an important concept
literature searches. All potentially relevant articles were for the analysis of networks, and betweenness centrality is
investigated as full text in English or Chinese. In cases of one of the most prominent measures of centrality. It is used
disagreement, a trial was included or excluded based on to measure the degree to which a node is in a position of
discussion between the two reviewers or after a third reviewer brokerage by summing up the fractions of shortest paths
reviewed the information. For duplicate publications, the between other pairs of vertices that pass through it [22].
final publication was used.
3. Results
2.5. Data Preprocessing. Information about titles, journals,
interventions, and main acupoints was extracted using the 3.1. Overall Profile of Acupuncture Prescriptions. Database
self-established Data Excavation Platform of Acupoint Speci- searching identified 892 records in CNKI, 1084 records in
ficity for data mining. Because acupoints have aliases, the CBM, and 145 records in PubMed. After screening, 238
names of acupoints were standardized according to Funda- acupuncture prescriptions in 238 articles were included.
mentals of Acupuncture [16]. Among the 238 trials, 185 are controlled clinical trials (CCTs),
while the other 53 trials have no controls. The whole view on
2.6. Data Processing the study quality of the 185 CCTs were shown in Figure 2.
2.6.1. Frequencies of Acupoints. The frequencies of acupoints, 3.2. Frequencies of Acupoints and Meridians. Approximately
meridians, and acupoints on different body parts were 109 meridian-acupoints distributed over 13 meridians and
analyzed using the Data Excavation Platform of Acupoint 7 extraordinary acupoints have been recorded for 1400 and
Specificity. 133 times, respectively, in modern literature on acupuncture
treatment for VaD. The most frequently used meridian
2.6.2.Association Rules Mining.Apriori Algorithm for associa- was the Governor Meridian (477 times). Other frequently
tion rules mining [17] was adopted to analyze the frequencies used meridians included the Gallbladder Meridian of Foot
Random sequence generation
Allocation concealment
Blinding of participants and personnel
Blinding of outcome assessment
Incomplete outcome data
Selective reporting
Another bias
0 20 40 60 80 100
(%)
Low risk
Unclear risk
High risk
Figure 2: Whole view on the study quality of the 185 CCTs.
200
180
160
140
120
100
80
60
40
20
0
Baihui (GV 20)
Sishencong (EX-HN 1)
Fengchi (GB 20)
Shuigou (GV 26)
Shenting (GV 24)
Neiguan (PC 6)
Zusanli (ST 36)
Sanyinjiao (SP 6)
Shenmen (HT 7)
Taixi (KI 3)
Benshen (GB 13)
Shenshu (BL 23)
Fenglong (ST 40)
Taichong (LV 3)
Dazhui (GV 14)
Xuanzhong (GB 39)
Fengfu (GV 16)
Hegu (LI 4)
Quchi (LI 11)
Yintang (EX-HN 3)
Frequency
Figure 3: The 20 most frequent acupoints and their frequencies.
Shaoyang and the Stomach Meridian of Foot Yangming, times), upper limbs (214 times), back and lumbar (127 times),
which were reported for 218 and 124 times, respectively. and chest and abdomen (43 times) (Figures 4(a) and 4(b)).
Extraordinary acupoints were also frequently used. Baihui
(GV 20), Sishencong (EX-HN 1), Fengchi (GB 20), Shuigou 3.4. Frequencies of Specific Acupoints. Specific acupoints rep-
(GV 26), and Shenting (GV 24), which were among the resented 78 of the 116 acupoints (67.24%). Specific acupoints
top five acupoints in frequency, were recorded for 176, 124, have been used 1292 times, representing 84.28% of the total
93, 86, and 84 times, respectively. The frequencies of each frequency of all acupoints (Figures 4(c) and 4(d)).
meridian and acupoint are shown in Table 1. The twenty most
frequently used acupoints are shown in Figure 3. 3.5. Frequencies and Support of Acupoint Combinations. Acu-
point combinations between Baihui (GV 20), Sishencong
3.3. Frequencies of Acupoints on Different Body Parts. Acu- (EX-HN 1), Fengchi (GB 20), Shenting (GV 24), Shuigou
points on the head, face, and neck were used most frequently, (GV 26), and Zusanli (ST 36) were used most frequently.
with a total number of 42 acupoints and a total frequency The 15 most frequently used acupoint combinations and their
of 766 times, followed by acupoints on the lower limbs (383 support and confidence are shown in Table 2.
Table 1: Statistics of meridians and acupoints in the modern literature on acupuncture treatment for VaD.
Number Meridian Frequency Number of acupoints Acupoints and their frequencies
Baihui (GV 20) 176, Shuigou (GV 26) 86, Shenting (GV 24) 84, Dazhui (GV 14) 27, Fengfu (GV 16) 23,
Yintang (EX-HN 3) 16, Naohu (GV 17) 13, Shangxing (GV 23) 11, Mingmen (GV 4) 8, Yamen (GV 15) 5,
1 GV 477 26 Qiangjian (GV 18) 5, Qianding (GV 21) 4, Yaoyangguan (GV 3) 3, Zhiyang (GV 9) 3, Jinsuo (GV 8) 2,
Shendao (GV 11) 1, Zhongshu (GV 7) 1, Taodao (GV 13) 1, Lingtai (GV 10) 1, Changqiang (GV 1) 1, Xuanshu
(GV 5) 1, Yaoshu (GV 2) 1, Shenzhu (GV 12) 1, Jizhong (GV 6) 1, Houding (GV 19) 1, Xinhui (GV 22) 1
Fengchi (GB 20) 93, Benshen (GB 13) 47, Xuanzhong (GB 39) 24, Shuaigu (GB 8) 7, Wangu (GB 12) 6,
Toulinqi (GB 15) 6, Qubin (GB 7) 5, Xuanli (GB 6) 4, Naokong (GB 19) 4, Zuqiaoyin (GB 44) 3, Yanglingquan
2 GB 218 20
(GB 34) 3, Yangbai (GB 14) 3, Hanyan (GB 4) 2, Qiuxu (GB 40) 2, Fengshi (GB 31) 2, Touqiaoyin (GB 11) 2,
Xuanlu (GB 5) 2, Muchuang (GB 16) 1, Zhengying (GB 17) 1, Zhongdu (GB 32) 1
Sishencong (EX-HN 1) 123, Taiyang (EX-HN 5) 4, Wailaogong (EX-UE 8) 2, Shiqizhui (EX-B 8) 1, Baxie
3 EX-HN 133 7
(EX-UE 9) 1, Anmian (EX-HN 22) 1, Yiming (EX-HN 14) 1
Zusanli (ST 36) 72, Fenglong (ST 40) 40, Touwei (ST 8) 6, Lidui (ST 45) 2, Sibai (ST 2) 2, Futu (ST 32) 1,
4 ST 124 7
Renying (ST 9) 1
Shenshu (BL 23) 43, Ganshu (BL 18) 12, Tianzhu (BL 10) 10, Pishu (BL 20) 8, Feiyang (BL 58) 8, Geshu (BL 17)
5 BL 103 15 7, Xinshu (BL 15) 4, Zhiyin (BL 67) 2, Kunlun (BL 60) 2, Yuzhen (BL 9) 2, Tongtian (BL 7) 1, Chengjin (BL 56)
1, Dazhu (BL 11) 1, Weizhong (BL 40) 1, Qucha (BL 4) 1
6 SP 96 5 Sanyinjiao (SP 6) 71, Xuehai (SP 10) 14, Taibai (SP 3) 8, Yinbai (SP 1) 2, Yinlingquan (SP 9) 1
7 PC 86 5 Neiguan (PC 6) 73, Zhongchong (PC 9) 4, Daling (PC 7) 4, Jianshi (PC 5) 3, Laogong (PC 8) 2
8 KI 83 5 Taixi (KI 3) 57, Dazhong (KI 4) 12, Yongquan (KI 1) 10, Zhaohai (KI 6) 8, Rangu (KI 2) 2
9 HT 67 3 Shenmen (HT 7) 62, Shaochong (HT 9) 3, Jiquan (HT 1) 2
Hegu (LI 4) 20, Quchi (LI 11) 18, Shangyang (LI 1) 2, Shousanli (LI 10) 2, Binao (LI 14) 1, Jianyu (LI 15) 1,
10 LI 45 7
Yingxiang (LI 20) 1
11 CV 43 5 Qihai (CV 6) 11, Zhongwan (CV 12) 10, Guanyuan (CV 4) 9, Danzhong (CV 17) 9, Shenque (CV 8) 4
12 LV 40 2 Taichong (LV 3) 37, Dadun (LV 1) 3
13 TE 11 5 Waiguan (TE 5) 6, Guanchong (TE 1) 2, Sizhukong (TE 23) 1, Jiaosun (TE 20) 1, Sidu (TE 9) 1
14 LU 4 2 Shaoshang (LU 11) 3, Lieque (LU 7) 1
15 SI 3 2 Shaoze (SI 1) 2, Yanggu (SI 5) 1
16 Total 1,533 116
GV, Governor Meridian; GB, Gallbladder Meridian of Foot Shaoyang; EX-HN, extraordinary acupoint; ST, Stomach Meridian of Foot Yangming; BL, Bladder Meridian of Foot Taiyang; SP, Spleen Meridian of Foot
Taiyin; PC, Pericardium Meridian of Hand Jueyin; KI, Kidney Meridian of Foot Shaoyin; HT, Heart Meridian of Hand Shaoyin; CV, Conception Vessel; LI, Large Intestine Meridian of Hand Yangming; LV, Liver
Meridian of Foot Jueyin; TE, Triple Energizer of Hand Shaoyang; LU, Lung Meridian of Hand Taiyin; SI, Small Intestine of Hand Taiyang. Frequencies of meridians refer to the total frequencies of acupoints on the
same meridian. Number of acupoints refer to the total number of acupoints on the same meridian.
5
Table 2: Statistics of the 15 most frequently used acupoint combinations in the treatment of VaD.
Number Acupoint combination Frequency Support (%)

1 Baihui (GV 20), Sishencong (EX-HN 1) 98 41.18
2 Baihui (GV 20), Fengchi (GB 20) 81 34.03
3 Baihui (GV 20), Shuigou (GV 26) 72 29.83
4 Baihui (GV 20), Shenting (GV 24) 70 29.41
5 Baihui (GV 20), Zusanli (ST 36) 62 26.05
6 Sishencong (EX-HN 1), Fengchi (GB 20) 60 25.21
7 Baihui (GV 20), Sanyinjiao (SP 6) 57 23.95
8 Baihui (GV 20), Neiguan (PC 6) 54 22.69
9 Sishencong (EX-HN 1), Shuigou (GV 26) 51 21.43
10 Sishencong (EX-HN 1), Baihui (GV 20), Fengchi (GB 20) 51 21.43
11 Baihui (GV 20), Shenmen (HT 7) 51 21.43
12 Sishencong (EX-HN 1), Shenting (GV 24) 49 20.59
13 Sishencong (EX-HN 1), Neiguan (PC 6) 46 19.33
14 Baihui (GV 20), Taixi (KI 3) 46 19.33
15 Sishencong (EX-HN 1), Baihui (GV 20), Shuigou (GV 26) 43 18.07
Support refers to the percentage of acupoint prescriptions containing the acupoint combination.
3.6. Community Structure. Community detection resulted (GV 24) are potential acupoints for treating VaD. In terms of
in the division of the 116 acupoints into 5 communities. meridian, acupoints on the Governor Meridian have poten-
Nodes of the same color belong to the same community. The tial for treating VaD. From the perspective of combinations,
community structure is shown in Figure 5(a). combinations between such acupoints as Baihui (GV 20),
Sishencong (EX-HN 1), Fengchi (GB 20), Shenting (GV 24),
3.7. Acupoint 𝐾-Core Network. The largest 𝑘-core value was Shuigou (GV 26), and Zusanli (ST 36) have potential for
19. At this value, there were 28 nodes, corresponding to Hegu treating VaD. In addition, acupoints on the head, face, and
(LI 4), Quchi (LI 11), Zusanli (ST 36), Fenglong (ST 40), neck have more potential for VaD than acupoints on other
Sanyinjiao (SP 6), Xuehai (SP 10), Shenmen (HT 7), Tianzhu regions of the body. Specific acupoints have more potential
(BL 10), Xinshu (BL 15), Ganshu (BL 18), Shenshu (BL 23), than nonspecific acupoints. Specific acupoints, with specific
Taixi (KI 3), Dazhong (KI 4), Neiguan (PC 6), Benshen (GB names, are a group of acupoints on fourteen meridians with
13), Fengchi (GB 20), Xuanzhong (GB 39), Taichong (LV 3), specific therapeutic effects. There are ten types of specific acu-
Mingmen (GV 4), Dazhui (GV 14), Yamen (GV 15), Fengfu points, Five-Shu acupoints, Yuan-Primary acupoints, Luo-
(GV 16), Baihui (GV 20), Shenting (GV 24), Shuigou (GV 26), Connecting acupoints, Xi-Cleft acupoints, Lower He-Sea
Guanyuan (CV 4), Sishencong (EX-HN 1), and Yintang (EX- acupoints, Back-Shu acupoints, Front-Mu acupoints, eight
HN 3), as shown in Figure 5(b).
influential acupoints, eight confluent acupoints connecting
the eight extra meridians, and convergent acupoints.
3.8. Degree. The top 20 acupoints in degree are shown
in Figure 6(a). Baihui (GV 20), Sishencong (EX-HN 1), Community detection divided the acupoints into 5 com-
Fengchi (GB 20), Shenting (GV 24), and Neiguan (PC 6) munities. Acupoints within the same community have some
had the highest degrees, with values of 89, 76, 68, 67, and 65, characteristics in common. Blue nodes (Community A) were
respectively. all Jing-Well acupoints. Yellow nodes (Community B) were all
acupoints on the face and head. Most green nodes (Commu-
3.9. Betweenness Centrality. The top 20 acupoints in between- nity C) were acupoints on the four limbs. Most purple nodes
ness centrality are shown in Figure 6(b). Yongquan (KI 1), (Community D) were acupoints belonging to Governor Ves-
Baihui (GV 20), and Sishencong (EX-HN 1) had the highest sel. Most red nodes (Community E) were specific acupoints
betweenness centrality. or acupoints with specific therapeutic effects, and only this
community contained Bach-Shu acupoints and acupoints
on the abdomen. Acupoints within the same community
4. Discussion were more densely connected with each other compared
4.1. Potential Acupoints and Combinations for VaD. In this with acupoints from different communities, indicating that
study, acupoints and combinations with potential for treat- an acupoint was more often used with acupoints within
ing VaD were discovered. These results may provide some the same community compared with acupoints within other
reference for the selection of acupoints in treatment for communities.
VaD, which may promote the therapeutic effects in clinical The 19-core network indicated that 28 acupoints, includ-
practice. The results suggest that Baihui (GV 20), Sishencong ing Hegu (LI 4), Quchi (LI 11), Zusanli (ST 36), Fenglong
(EX-HN 1), Fengchi (GB 20), Shuigou (GV 26), and Shenting (ST 40), Sanyinjiao (SP 6), Xuehai (SP 10), Shenmen (HT 7),
45 900
40 800
35 700
30 600
25 500
20 400
15 300
10 200
5 100
0 0
Head, face, Lower Upper Back and Chest and Head, face, Lower Upper Back and Chest and
and neck limbs limbs lumbar abdomen and neck limbs limbs lumbar abdomen
Number Frequency
(a) The numbers of acupoints in different parts (b) The frequencies of acupoints in different parts
40 800
35 700
30 600
25 500
20 400
15 300
10 200
5 100
0 0
Convergent acupoint
Five-Shu acupoint
Yuan-source point
Luo-Connecting point
Shu-Back point
Eight confluent acupoints
Front-Mu point
Lower He-Sea point
Nonspecifc point
Eight influential point
Convergent acupoint
Five-Shu acupoint
Yuan-source point
Luo-Connecting point
Shu-Back point
Eight confluent acupoints
Front-Mu point
Lower He-Sea point
Nonspecifc point
Eight influential point
Number Frequency
(c) The numbers of different kinds of acupoints (d) The frequencies of different kinds of acupoints
Figure 4: The frequencies and numbers of acupoints in different body parts and different types of acupoints.
Tianzhu (BL 10), Xinshu (BL 15), Ganshu (BL 18), Shenshu to 4 different communities, had higher betweenness cen-
(BL 23), Taixi (KI 3), Dazhong (KI 4), Neiguan (PC 6), trality. Yongquan (KI 1), which had the highest betweenness
Benshen (GB 13), Fengchi (GB 20), Xuanzhong (GB 39), centrality, did not have a relatively high degree. However, it
Taichong (LV 3), Mingmen (GV 4), Dazhui (GV 14), Yamen connects Jing-Well acupoints with other types of acupoints,
(GV 15), Fengfu (GV 16), Baihui (GV 20), Shenting (GV 24), resulting in a high betweenness centrality. Acupoints with
Shuigou (GV 26), Guanyuan (CV 4), Sishencong (EX-HN 1), higher betweenness centrality play an important role in
and Yintang (EX-HN 3) are core acupoints in the network. connecting different types of acupoints. Jing-Well acupoints,
Baihui (GV 20), Sishencong (EX-HN 1), Fengchi (GB except Yongquan (KI 1), were often used with other Jing-Well
20), Shenting (GV 24), and Neiguan (PC 6) had the highest acupoints only. Yongquan (KI 1) was not only used with other
degrees. This result indicates that these 5 acupoints have been Jing-Well acupoints but also with other types of acupoints,
combined with more acupoints than other acupoints. These such as other types of specific acupoints, nonspecific acu-
acupoints have specific therapeutic effects on VaD. Therefore, points, and acupoints on other parts. The high betweenness
these acupoints can be used together with other acupoints to centrality suggested that Yongquan (KI 1) may have multiple
enhance therapeutic effects. effects compared with other Jing-Well acupoints in treatment
Yongquan (KI 1), Baihui (GV 20), Sishencong (EX-HN of VaD. From the perspective of TCM theory, Jing-Well
1), Neiguan (PC 6), and Shenting (GV 24), which belonged acupoints can restore consciousness. Yongquan (KI 1) was
GV6
HT1
GV5 GV4
GV9 GV10
TE20
GV14
EX-B8 GV8 SP6
ST2 EX-HN22 BL56 EX-UE9
GV7 GV16
TE23
GV19 GV11
ST32 GB31 GV22
GV26 GV1 BL11
GV12 GB40 BL10
GV2
BL4 GB34
GV15 PC8 LI14
GV13
EX-HN14 GV20 TE9 BL40
GV3 GB32 GB16
CV6 BL60
SP9 ST36
GB39 GB17
LI11
CV8 CV12 LI10
BL18 LI4
LI15 TE5
ST9 LV3
PC6 PC7
EX-HN1 GB5
LU7 SI5 GB8
EX-HN5 BL9
SP3 EX-UE8 BL20 GB14
ST40
BL17 GB13 GV23
KI3 GB15
GV21
PC5 GV17
CV17 KI4 HT7 EX-HN3
CV4 BL23 ST8
GB20 KI6
GB19 GB7
BL15 SI1 LI20
BL58 SP1 GB6
SP10 GV24
GB12 BL7 GB4
KI1 TE1
KI2 LV1 ST45 GB11
GV18
HT9 LI1
GB44 PC9
BL67
LU11
(a) Network structure of acupoints for the treatment of VaD
GV4 KI4
GB13
EX-HN3
CV4
GV24 GV16
KI3
GV14
EX-HN1 GB39
ST40
BL23 GV20 PC6

GV26
SP6
GV15
HT7 LI4
GB20
ST36
BL18 LI11
LV3
BL15
BL10 SP10
(b) 19-core network
Figure 5: (a) Network structure of acupoints for the treatment of VaD. Blue nodes (Community A) are all Jing-Well acupoints. Yellow nodes
(Community B) are all acupoints on the face and head. Most green nodes (Community C) are acupoints on four limbs. Most purple nodes
(Community D) are acupoints belonging to Governor Vessel. Most red nodes (Community E) are specific acupoints or acupoints with
specific therapeutic effects, and only this community contains Bach-Shu acupoints and acupoints on the abdomen. Acupoints within the
same community are more densely connected with each other than acupoints from different communities. (b) 19-core network. There are 28
acupoints in the 19-core network. They are core acupoints in the treatment of VaD.
100
90
80
70
60
50
40
30
20
10
0
GV20
EX-HN1
GB20
GV24
PC6
SP6
GV14
GV26
GV16
HT7
LI4
KI3
ST36
ST40
LV3
GB13
EX-HN3
BL10
GB39
LI 11
Degree
(a) The 20 acupoints with highest degree
0.16
0.14
0.12
0.1
0.08
0.06
0.04
0.02
0
LV3
ST40
ST36
GB13
PC9
KI1
GV20
EX-HN1
PC6
GV26
GB20
GV24
GV14
SP6
EX-HN3
HT7
LI4
KI3
GV16
BL10
Betweenness centrality
(b) The 20 acupoints with highest betweenness centrality
Figure 6: The 20 acupoints with highest degree and the 20 acupoints with highest betweenness centrality.
also used to tonify kidney in treatment of VaD. Therefore, Electroacupuncture at Baihui (GV 20), Dazhui (GV 14), and
Yongquan (KI 1) is used not only with other Jing-Well Shenshu (BL 23) blocks expression of p53 and Noxa in the
acupoints but also with other types of acupoints, such as hippocampal CA1 region of VaD rats [26]. Acupuncture at
other types of specific acupoints, nonspecific acupoints, and Baihui (GV 20) can improve neurogenesis via regulating
acupoints on other parts. brain-derived neurotrophic factor (BDNF) and cyclic AMP
response element-binding protein (CREB). BDNF, which
4.2. Underlying Molecular Mechanism of the Acupoint with the is essential for synaptic plasticity and is coupled to CREB
Most Potential to Treat VaD. According to our results, Baihui activation [27], is important for long-term memory storage
(GV 20), which had the highest frequency, has the most [28]. CREB is required for the proliferation, growth, survival,
potential to treat VaD. A systematic review and meta-analysis and differentiation of all types of cells. In the brain, the CREB
also suggested that Baihui (GV 20) is a principal acupoint for and CRE-mediated system is involved in memory, learning,
acute intracerebral hemorrhage (ICH); in animal models of synaptic transmission, neuron survival, differentiation, and
acute ICH, there was no difference in efficacy between Baihui axon growth [29]. Acupuncture at Baihui (GV 20) signifi-
(GV 20) alone and Baihui (GV 20) plus other acupoints [23]. cantly increases the levels of BDNF [15, 30], CREB proteins,
Molecular biology studies have provided insights into CREB mRNA [30], and phosphorylated CREB, the active
the mechanisms underlying the effects of Baihui (GV 20) in form of CREB [15]. The molecular mechanism underlying
VaD treatment, including antioxidant effects, antiapoptotic acupuncture at Baihui (GV 20) also involves cholinergic
effects, neurotrophic effects, reduced blood-brain barrier system regulation. Decreased cholinergic function in the
(BBB) permeability, and regulation of the cholinergic and brain can result in a decline in memory and cognitive
dopaminergic systems. Acupuncture at Baihui (GV 20) in function [31]. Acupuncture at Baihui (GV 20) significantly
combination with other acupoints decreases levels of 8- increases the levels of choline acetyltransferase (ChAT) and
hydroxy-2󸀠 -deoxyguanosine, a product of oxidative damage restores the expression of choline transporter 1 (CHT1)
to DNA induced by free radicals, suggesting that the benefit and vesicular acetylcholine transporter (VAChT) [30]. The
of acupuncture is partly due to antioxidant effects [11]. dopaminergic system is also involved in the mechanism
Acupuncture exerts therapeutic effects on VaD via antiapop- underlying the treatment of VaD with acupuncture at Baihui
tosis. The tumor suppressors p53 and Noxa are important in (GV 20). Dopamine is a key regulator in specific synaptic
regulating apoptosis and mediate hypoxic cell death [24, 25]. changes observed at certain stages of learning and memory
and of synaptic plasticity [32]. Acupuncture at Baihui (GV 20) 4.4. Single Acupoint or Acupoint Combination. Acupoint com-
increases dopamine levels in chronic cerebral hypoperfusion binations also influence the therapeutic effects of acupunc-
and ischemia-reperfusion injured rats [33]. In addition, ture. An acupoint combination is considered to have a
acupuncture at Baihui (GV 20) and Zusanli (ST 36) preserves synergistic effect that enhances the therapeutic effect of
the integrity of the BBB, reducing BBB permeability. The acupuncture. For example, a lower prevalence of postoper-
BBB is constructed of tight conjunctions, including occludin ative nausea and vomiting in patients treated with Neiguan
and claudin-5, which form the endothelial barrier. Reduced (PC 6) plus Hegu (LI 4) was observed compared with those
expression of ZO-1, claudin-5, and occludin mRNA and treated with Neiguan (PC 6) only [42]. In spite of extensive
protein contributes to BBB breakdown and edema in the evidence suggesting a synergistic effect of acupoint combi-
ischemic brain [34]. Electroacupuncture at Baihui (GV 20) nation and supporting its use, some studies have reported
and Zusanli (ST 36) reduces brain damage and related antagonistic effects [43–45]. An antagonistic effect occurs
behavioral deficits via upregulation of tight conjunction when one acupoint weakens the therapeutic effect of another
proteins, including ZO-1, claudin-5, and occludin [35]. These acupoint [46]. For example, electroacupuncture can improve
findings reveal parts of the molecular mechanism underlying gastrointestinal movement in rats. The effect of needling
acupuncture at Baihui (GV 20) to treat VaD. Pishu (BL 20) alone was better than the effect of needling
Pishu (BL 20) and Zusanli (ST 36) at the same time [44].
Therefore, whether the effect of acupoint combination is
4.3. Acupoints Selection in Treatment for VaD. The proper better than a single acupoint still remains a question and
selection of acupoints is essential for the therapeutic effects needs to be further studied.
of acupuncture because acupoints are specific with regard to Some studies have compared single acupoints and an
morphological structure, biophysical properties, pathological acupoint combination for the treatment of VaD. The thera-
response, and stimulating effects [36]. This specificity dif- peutic effect of needling Baihui (GV 20), Shuigou (GV 26),
ferentiates acupoints from nonacupoints as well as different and Shenmen (HT 7) in combination was better than the
acupoints from one another. The specific therapeutic effects effects obtained by needling each alone [40]. In addition,
of different acupoints have been reported for migraine [37], needling Baihui (GV 20), Shuigou (GV 26), and Shenmen
functional dyspepsia [38], ischemic stroke [39], and so forth. (HT 7) simultaneously activated more brain areas related
The specificity of acupoints for the treatment of VaD to intellectual activities compared with needling each alone,
has also been reported. Phosphorylated CREB levels were generating a more extensive effect on the brain [41]. Antago-
significantly increased after acupuncture therapy of needling nistic effects in acupuncture therapy for VaD have not been
Baihui (GV 20) and Zusanli (ST 36) compared to sham reported but may occur. Most acupuncture prescriptions for
acupuncture therapy of needling nonacupoints [15]. Baihui VaD contain acupoint combinations, and the use of acupoint
(GV 20), Shuigou (GV 26), and Shenmen (HT 7) are all combinations is supported. However, acupoint combinations
among the 10 acupoints with the most potential. A clinical should be selected carefully to avoid antagonistic effects. It
trial demonstrated that needling Baihui (GV 20), Shuigou is hard to tell whether a combination of acupoints will exert
(GV 26), and Shenmen (HT 7) were all effective in improving antagonistic effects with current knowledge or TCM theory.
the symptoms of VaD. However, their therapeutic effects dif- As abovementioned, acupoints with similar functions can
fer. Needling Baihui (GV 20) improved calculation ability and exert antagonistic effects. There are many acupoints, and the
short-term memory and corrected the personality changes number of acupoint combinations will grow geometrically.
of VaD patients, while needling Shuigou (GV 26) improved To test the antagonistic effects of each combination one
naming ability and short-term memory. The therapeutic by one is an exhausting job. To avoid antagonistic effects
as possible, the acupoint prescriptions should be simplified
effects of needling Baihui (GV 20) and Shuigou (GV 26)
as possible. The general principle is to select acupoints
were superior to those of needling Shenmen (HT 7) [40].
with relatively better therapeutic effects and acupoints with
A PET and SPECT study revealed that needling these three
multiple indications and not to select many acupoints.
different acupoints in VaD patients affected different brain
areas. Needling Baihui (GV 20) activated the inner temporal
4.5. Limitations. This study has limitations as follows. First,
system, the thalamencephalon system, and the prefrontal other factors that influence acupuncture, such as manipula-
cortical system. Needling Shuigou (GV 26) activated the pre- tion and treatment duration, were not analyzed in this study.
frontal cortical system. Needling Shenmen (HT 7) generated These data can be further mined in future studies. Second,
an effect similar to but weaker than the effect generated by largely due to the lack of treatment based on syndrome differ-
needling Shuigou (GV 26) [41]. These findings demonstrate entiation and different methods of syndrome differentiation
that different acupoints have different therapeutic effects in in modern literature, potential acupoints and combinations
acupuncture treatment for VaD. Consequently, the selection for different syndromes of VaD were not analyzed. Although
of acupoints, which directly influences the therapeutic effects treatment based on syndrome differentiation is important
of acupuncture, should be considered carefully. According to and is often emphasized in TCM, treatment based on disease
our results based on data mining, Baihui (GV 20), Sishencong differentiation is equally important. Third, the real therapeu-
(EX-HN 1), Fengchi (GB 20), Shuigou (GV 26), and Shenting tic effects of acupoints and combinations on VaD cannot
(GV 24), which have higher frequencies in the modern be reflected by frequencies in the literature. However, these
literature, may have better therapeutic effects on VaD. results suggest some potential acupoints and combinations to
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The authors declare no conflict of interests. vascular dementia in 30 cases,” Journal of Traditional Chinese
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Authors’ Contribution ture protected cerebral multi-infarction rats from memory
Shuwei Feng and Yulan Ren contributed equally to this work. impairment by regulating the expression of apoptosis related
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This study was supported by the National Basic Research
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http://dx.doi.org/10.1155/2015/157897
Research Article
Effects of Deep Electroacupuncture Stimulation at
‘‘Huantiao’’ (GB 30) on Expression of Apoptosis-Related
Factors in Rats with Acute Sciatic Nerve Injury
Lili Dai,1,2 Yanjing Han,3 Tieming Ma,1 Yuli Liu,1 Lu Ren,1 Zenghua Bai,1 and Ye Li4
1
Liaoning University of Traditional Chinese Medicine, Shenyang 110847, China
2
Liaoning Health Vocational and Technical College, Shenyang 110101, China
3
4
Liaoning University of Traditional Chinese Medicine, Benxi Campus, Benxi 117000, China
Correspondence should be addressed to Tieming Ma; matieming999@sohu.com
Received 10 December 2014; Revised 30 January 2015; Accepted 5 February 2015
Academic Editor: Haifa Qiao
Copyright © 2015 Lili Dai et al. This is an open access article distributed under the Creative Commons Attribution License, which
SD rats were randomly divided into normal control, model, deep EA, and shallow EA groups. The model was established by
mechanical clamping of the sciatic nerve stem. For deep and shallow EA, the needles were inserted into “Huantiao” (GB 30) by
about 16 mm and 7 mm, respectively, once daily for 14 days. The results showed that, compared with the normal control group,
the nerve-muscle excitability of rat’s hip muscle decreased and the nerve conduction velocity of sciatic nerve slowed down in the
model group; meanwhile, the number of apoptotic cells and the expression level of Bax protein in the injured nerve increased
significantly, and the expression level of Bcl-2 protein and the ratio of Bcl-2/Bax decreased considerably. Compared with the model
group, the indices mentioned above were reversed in the two treatment groups, and the changes in the deep EA group were more
significant than those in the shallow EA group. These results indicate that EA stimulation at GB 30 can improve the function of
injured sciatic nerve, which is closely associated with its effects in upregulating the expression of apoptosis inhibitive factor Bcl-2
and downregulating apoptosis promotive factor Bax. Deep EA is relatively better.
1. Introduction practice confirmed that the sense of electric shock induced

by deep acupuncture at Huantiao (GB 30) had a significant
Peripheral nerve injury is one of the most common traumatic effect on the functional recovery of injured nerves [8, 9].
disorders. Slow recovery and prolonged loss of sensation or Experimental studies showed that acupuncture at GB 30 had
motor function may cause muscle atrophy, joint contracture, a positive influence on motor recovery [4] and facilitated
and deformity. If the injured nerve fibers are repaired, the axonal regeneration in the injured peripheral nerves [10].
continuity of nerves can be restored, providing favorable Previous work by our research group indicated that deep
conditions for nerve regeneration. electroacupuncture (EA) stimulation at GB 30 improved
Acupuncture has been proven to be an effective method the pathological changes and function of the injured sciatic
for the treatment of peripheral nerve injury and is widely used nerve in rats, which was closely associated with its effects
to promote the recovery of nerve function [1, 2]. Research on the upregulation of nerve growth factor expression and
has shown that the nerve stump is ischemic in the early downregulation of Fos expression in the damaged sciatic
stage and that local blood circulation can be improved by nerve. Deep EA was found to be better than shallow EA [11].
acupuncture therapy [3]. With regard to the related mecha- The present study aimed to further investigate the mechanism
nism, research has proved the positive effect of acupuncture of repair of sciatic nerve injury following acupuncture at GB
on the repair of injured nerves from the perspectives of 30 and the difference between deep EA and shallow EA in
behavior, electrophysiology, and morphology [4–7]. Clinical terms of apoptosis.
2. Materials and Methods (the depression in front of the femoral greater trochanter at
the leading edge of the hip joint in the affected side) with the
2.1. Animals and Grouping. This experiment was conducted depth of about 16 mm to the extent that the muscle twitched
in accordance with the Guide for Care and Use of Laboratory instantly and the toes trembled. The electroacupuncture ther-
Animals issued by the National Institutes of Health. apeutic apparatus was then connected, while the indifferent
Forty-eight healthy pathogen-free Sprague Dawley (SD) electrode was placed in the homolateral lower limb. Dilata-
rats (24 male and 24 female) with a body mass of 250 ± tional wave was applied with a frequency of 2 Hz/100 Hz and
20 g, were provided by the Experimental Animal Center, an intensity of about 2 mA and the affected limb was observed
Liaoning University of Traditional Chinese Medicine, license to twitch slightly. The treatment was given for 20 min each
number: SCXK (Liaoning) 2008-0005. The laboratory envi- time, once daily for 14 days.
ronment was as follows: temperature was 18–22∘ C, indoor
light exposure was approximately 8 h, and relative humidity Shallow EA Group. The sciatic nerve injury model was
was about 45%. Free access to water and food was allowed, achieved by mechanical clamping of the sciatic nerve stem.
and males and females were kept separately with six rats per After successful modeling, acupuncture was applied to the
cage. According to the random number table, the rats were GB 30 with the depth of about 7 mm without touching the
randomly divided into the normal control, model, deep EA, nerve trunk. The electroacupuncture therapeutic apparatus
and shallow EA groups with 12 in each group. was then connected, while the indifferent electrode was
placed in the homolateral lower limb. Dilatational wave was
2.2. Main Reagents and Instruments. The following reagents applied with a frequency of 2 Hz/100 Hz and an intensity of
and instruments were used: cell apoptosis detection kit about 2 mA and the affected limb was observed to twitch
(Roche), Bcl-2 (B-cell lymphoma/leukemia-2), and Bax (Bcl- slightly. The treatment was given for 20 min each time, once
2 associated X protein) immunohistochemical detection kits daily for 14 days.
(Shanghai BlueGene Biotech Co., Ltd.), Hwato acupunc-
ture needles (Suzhou Acupuncture Supplies Factory), pulse 2.5. Criteria of Successful Modeling. Half an hour after mod-
electroacupuncture therapeutic apparatus (6805-A, Shantou eling, 5 rats were randomly selected for EMG testing to
Medical Equipment Factory), biophysiological experimental observe the motor conduction velocity (MCV) of sciatic
system (BL-420, Chengdu Taimeng Electronics Co., Ltd.), nerves. When the MCV dropped below 10 m/s, it was deemed
microtome (RM 2235, Leica), digital microscope (BX 41, that the sciatic neuraxon and myelin sheath had broken or
Olympus), and MetaMorph microscopic image analysis sys- were severely injured [13].
tem (UIC, Olympus).
2.6. Observation Indices and Methods
2.3. Modeling. The acute sciatic nerve injury model was
established by mechanical clamping of the sciatic nerve General Status. The rats’ mental state, limb activity, reaction,
stem [12]. The rat was placed in the prone position on the ingestion, water intake, and daily activities were assessed.
operating table, and anesthesia was given by intraperitoneal
injection of 1% pentobarbital sodium (40 mg/kg). Routine Determination of the Strength-Duration (S-D) Curve. The S-
skin preparation and sterilization were carried out, and a 1 cm D curve was determined by applying the biophysiological
vertical incision was made at the rear of the middle femoral experimental system to display the nerve-muscle excitability
shaft on the left to expose the bicep femoris muscle. The in the rat. The working electrode was placed on the buttock,
sciatic nerve was then dissociated by blunt dissection and and the auxiliary electrode was placed on the ankle joint
then clamped with a 16 cm needle holder 0.5 cm below the of the homolateral posterior limb. First, the motor point of
femoral tubercle. The holder was released after squeezing for the muscle was detected by strong stimulation, and then
10 s. This was repeated 3 times with an interval of 10 s until the the magnitude of the current was turned down when weak
sciatic nerve was seriously injured. The sciatic nerve with the muscle twitches were observed with the naked eye. The
injured trunk of about 3 mm was marked by a 9-0 noninvasive stimulus threshold was measured with a pulse width of
suture thread and then put back in place, and the skin was 0.1–1 ms, and the S-D curve was drawn using logarithmic
sutured. The above operation was performed by one person. coordinates.
Detection of Conduction Velocity of the Sciatic Nerve. The rat

2.4. Processing Methods for Each Group was placed in the prone position on the operating table, and
anesthesia was given by intraperitoneal injection of 10% chlo-
Normal Control Group. The rats were kept under the same
ral hydrate (0.35 mg/100 g). The skin and muscles were cut
conditions without modeling and treatment.
using the modeling method to fully expose the sciatic nerve
Model Group. The rats were kept under the same conditions segment for surgery. The sciatic nerve was dissociated using
after modeling without any treatment. a glass dissecting needle with two insulated bipolar acicular
electrodes hooked at both ends of the nerve anastomosis,
Deep EA Group. The sciatic nerve injury model was achieved and the recording electrode was placed at the distal end
by mechanical clamping of the sciatic nerve stem. After of the stimulation electrode. The stimulus was then applied
successful modeling, acupuncture was applied to the GB 30 to determine the threshold causing evoked action potential.
This stimulus was repeated until the graph of action potential 1.2
Intensity of electric current (V)

on the screen remained stable and the starting points of the 1.1
artifact and action potential became clear. Screenshots were
1.0 ∗∗
then obtained, and the amplitude of action potential of the
nerve trunk was automatically displayed. If the amplitude 0.9
of action potential of the nerve trunk was measured from 0.8
the highest point to the lowest point, the measurement of Δ
0.7
the latent period lasted from the appearance of stimulus
0.6
artifact to the initiation site of action potential. When the
distance between the two stimulation electrodes was input, 0.5
0 0.1 0.2 0.3 0.4 0.5 0.6 0.7 0.8 0.9 1.0
the nerve conduction velocity was automatically displayed.
Pulse width (ms)
During the operation, the sciatic nerves were covered with
saline-soaked gauze to ensure that the exposed nerves and Normal control group Shallow EA group
muscles remained moist. Deep EA group Model group
Detection of Cell Apoptosis by Terminal Deoxynucleotidyltrans- Figure 1: Effect of deep and shallow electroacupuncture (EA)
stimulation of the ipsilateral “Huantiao” (GB 30) on the S-D curve
ferase-Mediated dUTP-Biotin Nick-End Labeling (TUNEL). 8
in hip muscles of the rats with injured sciatic nerves (𝑥 ± 𝑠, 12
rats randomly selected in each group were perfused with
rats/group). ∗∗ 𝑃 < 0.01 versus the normal control group; Δ 𝑃 < 0.05
4% paraformaldehyde and fixed on the operating table. The versus the model group.
injured sciatic nerve tissues were removed and fixed for
24 h. Routine paraffin sections (5 𝜇m) were prepared. After
being dewaxed, the sections were treated according to the
TUNEL kit instructions. Two sections were taken out from decreased with prolonged stimulus time; the lower segment
each rat. Eight nonoverlapped views were randomly selected (0.6–1 ms) appeared flat, indicating that stimulus intensity
for each section. The MetaMorph microscopic image analysis remained constant when stimulus time was long. Under a
system was applied. Brown yellow particles in the nucleus certain stimulus time, the voltage which aroused excitability
were positive cells, and the number of apoptotic cells was of the sciatic nerve in the four groups was the greatest in the
measured. model group, followed by the shallow EA group, the deep
EA group, and the normal control group. Compared with
Determination of the Expression of Bcl-2 and Bax Protein the normal control group, the voltage in the model group
by Immunohistochemistry. After being dewaxed, the sections increased significantly (𝑃 < 0.01); compared with the model
were treated with streptavidin-peroxidase (SP) immunohis- group, the voltage reduced significantly in the deep EA group
tochemistry. Three sections were taken out from each rat. (𝑃 < 0.05), which showed that after two weeks of treatment,
Five no-overlapped views were randomly selected for each the best recovery of injured sciatic nerve was achieved in the
section. The MetaMorph microscopic image analysis system deep EA group. The second best recovery of injured sciatic
was used to measure the mean optical density of the positive nerve occurred in the shallow EA group.
products of Bcl-2 and Bax. The ratio of Bcl-2/Bax was then
calculated. 3.3. Changes in Conduction Velocity of Sciatic Nerves in the
Four Groups. As shown in Figure 2, the conduction velocity
2.7. Statistical Analysis. SPSS 17.0 statistical software was used in the model group decreased (𝑃 < 0.01), suggesting
to analyze the data, and the results are presented as mean ± segmental demyelination of most nerve fibers; compared with
standard deviation (𝑥 ± 𝑠). Variance was applied to evalu- the model group, the conduction velocity in the deep and
ate integral differences, and equal variance was compared shallow EA groups increased significantly (𝑃 < 0.05); the
between the two groups by means of LSD. A nonparametric conduction velocity in the shallow EA group was markedly
test was used in the case of nonconformity with normal lower than that in the deep EA group (𝑃 < 0.05).
distribution. 𝑃 < 0.05 was considered statistically significant.
3.4. Changes in the Number of Apoptotic Cells in the Injured
3. Results Sciatic Nerves in the Four Groups. As shown in Figure 3, the
number of apoptotic cells in the model group was increased
3.1. Observation of General Status. After modeling, the intake compared with that in the normal control group (𝑃 < 0.05),
of food and water and defecation in the rats were normal. while the numbers of apoptotic cells in the deep EA and
Infection and ulcers were not found on the distal limb. The shallow EA groups were significantly decreased compared
rats walked by dragging toes or bouncing. Gait began to with those in the model group (𝑃 < 0.05). The effect of deep
recover one week after treatment in the deep EA and shallow EA was better than that of shallow EA (𝑃 < 0.05).
EA groups.
3.5. Changes in the Expression of Bcl-2 and Bax in Injured Sci-
3.2. Changes in the S-D Curve in the Four Groups. As shown atic Nerves in the Four Groups. As shown in Figures 4 and 5,
in Figure 1, the upper segment of the S-D curve (0.1–0.5 ms) the positive immunoreactivity of Bcl-2 and Bax proteins in
in each group was steeper, indicating that stimulus intensity the Schwann cell cytoplasm appeared brown. In the model
10.0 10.0
8.0 8.0
6.0 6.0 60
4.0 4.0
2.0 2.0
Sciatic nerve conduction velocity (m/s)

(mV)
50
(mV)
0 0
−2.0 −2.0
−4.0 −4.0
−6.0 40 Δ#
−6.0
−8.0 −8.0
−10.0 1.000 V 1.000 V −10.0 1.000 V
30 Δ
0 10 30 50 70 0 10 30 50
(ms) (ms)
(a) (b) 20
10.0
8.0 8.0 ∗∗
6.0 6.0
4.0 10
4.0
2.0
(mV)
2.0
0 0
(mV)
−2.0 −2.0
−4.0 0
−4.0 Normal control Model Deep EA Shallow EA
−6.0 −6.0
−8.0 −8.0
−10.0 1.000 V −10.0 1.000 V
1.000 V 1.000 V
0 10 30 50 70 90 0 10 30 50
(ms) (ms)
(c) (d)
Figure 2: Effect of deep and shallow EA stimulation of ipsilateral GB 30 on the conduction velocity of the injured sciatic nerve in rats (𝑥±𝑠, 12
rats/group). (a) Normal control group, (b) model group, (c) deep EA group, and (d) shallow EA group. ∗∗ 𝑃 < 0.01 versus the normal control
group; Δ 𝑃 < 0.05 versus the model group; # 𝑃 < 0.05 versus the shallow EA group.
60
∗
50
Number of apoptotic cells
Δ
40 *
(a) (b) 30 Δ#
20
10
0
Normal control Model Deep EA Shallow EA
(c) (d)
Figure 3: Effect of EA stimulation of ipsilateral GB 30 on number of apoptotic cells in the injured sciatic nerve in rats. Left panel: photos of
TUNEL staining showing the number of apoptotic cells in the sciatic nerve (indicated by black arrowheads) in the normal control (a), model
(b), deep EA (c), and shallow EA (d) groups (×200); right panel: bar graphs showing the number of apoptotic cells in the sciatic nerve in the
four groups (𝑥 ± 𝑠, 8 rats/group). ∗ 𝑃 < 0.05 versus the normal control group; Δ 𝑃 < 0.05 versus the model group; # 𝑃 < 0.05 versus the shallow
EA group.
0.35
0.30
Δ#
Mean optical density

0.25
Δ
0.20
(a) (b) 0.15

∗
0.10
0.05
0.00
(c) (d)
Figure 4: Effect of EA stimulation of ipsilateral GB 30 on Bcl-2 immunoreactivity in the injured sciatic nerve in rats. Left panel: photos of
immunohistochemical staining showing the expression of Bcl-2 in the sciatic nerve (indicated by red arrowheads) in the normal control (a),
model (b), deep EA (c), and shallow EA (d) groups (×200); right panel: bar graphs showing the expression levels (OD values) of Bcl-2 in the
sciatic nerve in the four groups (𝑥 ± 𝑠, 8 rats/group). ∗ 𝑃 < 0.05 versus the normal control group; Δ 𝑃 < 0.05 versus the model group; # 𝑃 < 0.05
versus the shallow EA group.
0.35
∗
0.30
Δ
0.25
Mean optical density
Δ#
0.20
(a) (b) 0.15
0.10
0.05
0.00
(c) (d)
Figure 5: Effect of EA stimulation of ipsilateral GB 30 on Bax immunoreactivity in the injured sciatic nerve in rats. Left panel: photos of
immunohistochemical staining showing the expression of Bax in the sciatic nerve (indicated by red arrowheads) in the normal control (a),
model (b), deep EA (c), and shallow EA (d) groups (×200); right panel: bar graphs showing the expression levels (OD values) of Bax in the
sciatic nerve in the four groups (𝑥 ± 𝑠, 8 rats/group). ∗ 𝑃 < 0.05 versus the normal control group; Δ 𝑃 < 0.05 versus the model group; # 𝑃 < 0.05
versus the shallow EA group.
group, the expression of Bcl-2 in sciatic nerve was signifi- Bcl-2/Bax in the model group was markedly lower than that
cantly lower than that in the normal control group, while in the normal control group (𝑃 < 0.05), while the ratios in
the expression of Bax was significantly higher (𝑃 < 0.05); the deep EA and shallow EA groups were significantly higher
compared with the model group, the expression of Bcl-2 than those in the model group (𝑃 < 0.05). The ratio in the
increased and that of Bax decreased in the deep EA and deep EA group was higher than that in the shallow EA group
shallow EA groups (𝑃 < 0.05). The difference between the (𝑃 < 0.05).
deep EA group and the shallow EA group was statistically
significant (𝑃 < 0.05).
4. Discussion
3.6. Changes in the Ratio of Bcl-2/Bax in the Injured Sciatic This study was designed to investigate the biological mech-
Nerves in the Four Groups. As shown in Figure 6, the ratio of anism involved in the differences in repair of sciatic nerve
3.50
3.00
Ratio of Bcl-2/Bax
2.50
2.00
1.50 Δ#
1.00 Δ
0.50 ∗
0.00
Figure 6: Effect of deep and shallow EA stimulation of ipsilateral GB 30 on the ratio of Bcl-2/Bax of the rats with injured sciatic nerves (𝑥 ± 𝑠,
8 rats/group). ∗ 𝑃 < 0.05 versus the normal control group; Δ 𝑃 < 0.05 versus the model group; # 𝑃 < 0.05 versus the shallow EA group.
injury by deep EA and shallow EA at GB 30. The results different pulse width. It is reflected by the curve of threshold
showed that, compared with shallow EA, deep EA at GB of current strength when the muscle is excited. The lighter
30, when the nerve trunk was reached, had a significantly the nerve injury, the less denervation, and the lower the S-D
better effect on the recovery of injured sciatic nerve. Deep curve, the lower the stimulus intensity to cause nerve-muscle
EA improved the excitability of the nerve and promoted the excitation. The results of this study showed that in the deep
recovery of motor nerve conduction, which may have been EA group, the current intensity to induce nerve excitement
achieved by the increased expression of Bcl-2 and reduced was markedly lower than that in the model group, indicating
expression of Bax, resulting in fewer apoptotic cells. better efficacy of deep EA in promoting the recovery of
Bcl-2 gene families play important roles in apoptosis. injured sciatic nerve. As a direct manifestation of nerve
According to their different roles, they are divided into two impulse conduction, the conduction velocity can reflect the
categories, one category can promote cell apoptosis; the other regeneration conditions of nerve fibers to some extent. The
can inhibit cell apoptosis. Bcl-2 and Bax of the Bcl-2 gene nerve trunk cannot be completely destroyed by clamping
families are two protein expression products closely related to sciatic nerves. Therefore, the rats, in which nerve conduction
apoptosis [14]. Bcl-2 is a type of membrane stabilizing protein was completely blocked and the electric waves could not be
related to organelles, especially mitochondria, and is mainly displayed, were excluded from this study.
found in the outer membrane of mitochondria, endoplasmic The treatment of flaccidity syndrome by acupuncture at
reticulum membrane, and nuclear membrane. Inhibition of GB 30 can be traced back to the Internal Canon of Medicine.
apoptosis can be achieved by inhibiting permeability of the The depth of acupuncture may determine its clinical efficacy
mitochondrial membrane, maintaining the stability of the [28, 29]. In recent years, the depth of acupuncture at GB
membrane, preventing the release of cytochrome C, and 30 has been increased. It is proposed that 2-3 cun is appro-
inhibiting the activation of Caspase by inhibition of free priate to ensure that the nerve trunk is reached [30–32].
radical generation and intracellular calcium overload caused Studies have shown that deep acupuncture at GB 30 may
by Bcl-2. Cell damage can be reduced by the overexpression have immediate and remarkable therapeutic and analgesic
of Bcl-2 [15–17]. In contrast, the biological effect of Bax effects on sciatic nerve injury [33]. Research on the related
differs from that of Bcl-2 and can promote apoptosis [18, mechanisms suggests that, for sciatic nerve injury in rabbits,
19]. Bcl-2 and Bax may interact as a dimer. When Bcl- acupuncture therapy can increase the content of acetylcholine
2 increases, the Bcl-2 homodimer is formed, and cells are esterase (AchE) in the intumescentia lumbalis of the spinal
protected; when Bax increases, the Bax/Bcl-2 heterodimer is cord. By promoting the synthesis and release of AchE, nerve
formed, and cell apoptosis occurs [20–22]. Thus, the ratio of excitability was maintained thus promoting the recovery of
Bcl-2/Bax directly determines cell survival. The number of nerve function [34]. EA can increase the content of 1L-
apoptotic cells is the balanced result of the two regulation 1𝛽 in spinal cord tissue and stimulate the production of
factors, Bcl-2 and Bax [23]. It is generally recognized that nerve growth factors (NGFs) [35], thus strengthening the
the main form of neuronal cell death caused by peripheral transcription of NGF mRNA and exerting a positive effect
nerve injury is apoptosis, a process of programmed cell on the repair of injured sciatic nerves [36]. The results of this
death involving a variety of physiological and pathological study showed that, compared with shallow EA, deep EA at
factors and initiated by apoptosis-related genes [24–26]. GB 30 better adjusted the substances related to the regulation
Kotulska et al. [27] suggested that the expression of Bcl-2 and of apoptosis, thus significantly improved the excitability and
Bax is closely related to the recovery and viability of neurons conduction of the sciatic nerve.
after peripheral nerve injury as well as fiber regeneration and
myelination. In this study, the function of the sciatic nerve in 5. Conclusion
rats in each group varied due to changes in Bcl-2 and Bax at
the injured site, which is consistent with previous results. Better functional recovery of injured sciatic nerve may be
The S-D curve is used to evaluate the function of inner- achieved by acupuncture at GB 30 when the nerve trunk is
vation by stimulating muscle with a square wave current of reached. By stimulating the axis cylinder with continuous
and chronic demyelination, strong stimulation is generated, [11] Y. L. Liu, Y. Li, L. Ren et al., “Effect of deep electroacupuncture
triggering strong nerve impulses to transfer nerve substances, stimulation of Huantiao (GB 30) on changes of function and
which can inhibit the apoptosis of nerve cells in the injured nerve growth factor expression of the injured sciatic nerve in
area and promote tissue repair. rats,” Acupuncture Research, vol. 39, no. 1, pp. 93–99, 2014.
[12] M. Sendtner, H. Schmalbruch, K. A. Stockli, P. Carroll, G.
W. Kreutzberg, and H. Thoenen, “Ciliary neurotrophic factor
Conflict of Interests prevents degeneration of motor neurons in mouse mutant
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[13] M. S. Alrashdan, J.-C. Park, M.-A. Sung et al., “Thirty minutes
Authors’ Contribution of low intensity electrical stimulation promotes nerve regen-
eration after sciatic nerve crush injury in a rat model,” Acta
The first two authors Lili Dai and Yanjing Han contributed Neurologica Belgica, vol. 110, no. 2, pp. 168–179, 2010.
equally to this work. [14] O. Nesic-Taylor, D. Cittelly, Z. Ye et al., “Exogenous Bcl-XL
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This work was supported by Natural Science Foundation of of Bcl-2, Bcl-xl, and Bax proteins after sublethal forebrain
ischemia in C57Black/Crj6 mouse striatum,” Stroke, vol. 34, no.
Liaoning Province (no. 201102146).
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http://dx.doi.org/10.1155/2015/516851
Research Article
Function of Nucleus Ventralis Posterior Lateralis
Thalami in Acupoint Sensitization Phenomena
Pei-Jing Rong,1 Jing-Jun Zhao,1 Ling-Ling Yu,1,2 Liang Li,1 Hui Ben,1
Shao-Yuan Li,1 and Bing Zhu1
1
Institute of Acupuncture and Moxibustion, China Academy of Chinese Medical Sciences, 16 Nanxiaojie of Dongzhimen,
2
Wuhan Integrated TCM and Western Medicine Hospital, 215 Zhongshan Road, Qiaokou District, Wuhan, Hubei 43002, China
Correspondence should be addressed to Bing Zhu; zhubing@mail.cintcm.ac.cn
Received 20 October 2014; Accepted 11 December 2014
Academic Editor: Jang-Hern Lee
Copyright © 2015 Pei-Jing Rong et al. This is an open access article distributed under the Creative Commons Attribution License,
To observe the effect of electroacupuncture (EA) on nucleus ventralis posterior lateralis (VPL) thalami activated by visceral noxious
stimulation and to explore the impact of EA on the mechanism of acupoint sensitization under a pathological state of the viscera,
EA was applied at bilateral “Zusanli-Shangjuxu” acupoints. The discharge of VPL neurons was response to EA increased after
colorectal distension (CRD). The stimulation at “Zusanli-Shangjuxu” acupoints enhanced discharge activity of VPL neurons under
CRD-induced visceral pain. The frequency of neuronal discharge was associated with the pressure gradient of CRD which showed
that visceral noxious stimulation may intensify the body’s functional response to stimulation at acupoints.
1. Introduction observed whether the effect of acupuncture on the receptive

field (acupoint area) of VPL neurons on body surface was
Acupoints are special locations on body surfaces where the Qi affected by visceral noxious inputs. The phenomena and
of meridians and internal organs is infused. They are also the mechanism of acupoint sensitization at the VPL level induced
key link underlying the interactions between meridians and by visceral noxious inputs will be discussed.
internal organs. When internal organs are under a pathologi-
cal state, acupoints become more sensitive [1–3]. The size and
function of acupoints change accordingly with the change of 2. Materials and Methods
visceral functions [4, 5]. Therefore in pathological conditions,
2.1. Experimental Animals. Twenty-six healthy male Spra-
the diagnostic and therapeutic effects of acupoints on visceral
diseases are enhanced [6]. gue-Dawley rats weighing 250–300 g were provided by the
Spinothalamic tract is traditionally viewed as the major Laboratory Animal Center of Academy of Military Medical
pathway of noxious inputs. Previous studies showed [7–10] Sciences (animal certificate Lot Scxr (Beijing) 2009-0017).
that noxious inputs transmitted via spinothalamic tract can Before experiments, the animals were fasted for 12 hours but
be affected by other noxious inputs. A current issue in neuro- they were not deprived of water. Throughout the experiment,
science research is the mechanism underlying the peripheral the animals’ body temperature was maintained between 36
and central sensitization caused by different noxious inputs and 38∘ C by a temperature control device (Model: CL-8;
[11]. Manufacturer: RWD/China). Animal experimental methods,
This study evaluated the neuronal discharge of ventral experimental purposes, and the disposal of animals in exper-
posterior lateral nucleus (VPL; the most important brain iment followed the Guidelines on Proper Care of Experimental
center for somatovisceral relay) by noxious inputs from Animals promulgated in 2006 by the Ministry of Science and
the body surface and colorectal distension (CRD). We also Technology.
2.2. Experimental Methods

Sphygmomanometer
2.2.1. Recording Discharge of Thalamic VPL Neurons. Rats
were anesthetized with an intraperitoneal injection of 10%
Computer
BIOPAC
urethane (1.0∼1.2 g/kg, provided by Shanghai Sinopharm Calibration MP150 TSD104A
Chemical Reagent Co., Ltd.). The heads of rats were fixed
on stereotaxic instruments. We incised the skin of the
middle of skull and the suture was exposed by removing Pressure
transducer
the resubcutaneous tissue and periosteum. Then we should
adjust the frontal and back suture located in a horizontal Figure 1: Experimental layout of colorectal distension.
plane. The three-dimensional location coordinates of VPL
nuclei were determined according to Rat Brain Atlas [12]
3.0∼4.0 mm behind the anterior fontanel, 3.0∼3.5 mm next to 0 BG 15󳰀 0 EA 30 0 CRD 30 0 EA 30 s
skull sutures. Under observation with a surgical microscope, Rest 10󳰀 Rest 10󳰀
the tip of the glass microelectrode was inserted to VPL nuclei
through the skull hole by the microelectrode manipulator Figure 2: Experimental flow chart.
(5000∼5800 𝜇m beneath the surface of the brain). Impedance
at the tip of the glass microelectrode was set at 10–15 MΩ
(filled with 2% pontamine sky blue). When the target neurons 2.3. Experimental Procedure. (1) The background discharge
were identified, 2% agar was perfused onto the skull surface of convergent neurons was recorded for 10–15 min using
to protect the brain tissue from drying and reduce volatility microelectrode amplifier (Model: MEZ 8201; Manufacturer:
caused by breathing. Nihon Kohden) and biological signal acquisition and anal-
For all recorded neurons, the responses to mechanical ysis system (Model: MICRO 1401; Manufacturer: British
stimulations applied to their peripheral receptive field were CED Company). (2) EA was applied at bilateral “Zusanli-
checked to identify the distribution and size of the receptive Shangjuxu” points for 30 seconds. (3) After an interval of 10
fields (mechanical stimulations include touch and pressure minutes, different intensities of CRD were given to rats for
by von Frey hairs (von Frey Model 2390; U.S. IITC Com- 30 seconds. The discharge of the convergent neurons to non-
pany), skin stimulation by tooth tweezers, and acupuncture noxious stimuli (20 mmHg), noxious stimuli (40 mmHg),
stimulation). We also observed responses of these neurons to and strongly noxious stimuli (60, 80 mmHg) was recorded,
CRD. Only neurons that responded to both the mechanical respectively, to observe the activation of convergent neurons
stimulation on the skin receptive field and the 10 mmHg of by different intensities of CRD. (4) After an interval of 10
CRD were included as the objects of observation (and were minutes, EA was once again applied at bilateral “Zusanli-
named as convergent neurons or CN). Shangjuxu” points for 30 seconds. The discharge of VPL neu-
rons in response to EA stimulation before and after different
2.2.2. Colorectal Distension. A 4 to 6 cm long balloon was intensities of CDR was observed to test the dose-effect rela-
made from a disposable condom tip and tied on a 4 mm tionship between stimulus intensity and response (Figure 2).
diameter hose (Figure 1; BIOPAC Amplifier Module Model:
MP150 System TSD104A; Manufacturer: BIOPAC Company, 2.4. Statistical Analysis. The data was analyzed with Spike-II
USA). The balloon was inserted through the anal orifice (the data analysis software of MICRO 1401 biological signal
straight into the colon. The depth was approximately 4 cm. acquisition and analysis system) and SPSS 13.0 software. The
Three to five drops of the warmed paraffin oil were smeared number of neuronal discharge of VPL neurons in every 30
on the balloon’s surface before the balloon was placed into seconds and the activation/inhibition rate were counted and
the colon to avoid direct damage to the inner wall of the the mean and variance of neuronal discharge before and after
colon and anus. The distance from balloon end to anus was the EA intervention were calculated. Comparison between
about 0.5 cm. 20–80 mmHg CRD stimulus was given via a groups was made with independent sample 𝑡-test. 𝑃 < 0.05
syringe, with the duration of about 30 s. The activation of was considered as statistically significant.
convergent neurons was observed at different intensities of
CRD stimulation. Pressure ≥ 40 mmHg was identified as 2.5. Histological Localization. When recording of neuronal
visceral noxious stimulation [13]. The time interval between discharge was completed, 20 𝜇A of negative direct current
CRD stimulations was no less than 10 min to avoid colorectal was passed to the glass microelectrode via the microelectrode
sensitization caused by hyper stimulation. amplifier for 20–30 min. Pontamine sky blue in the glass
microelectrode was imported into VPL nuclei to mark the
2.2.3. EA. EA was applied at bilateral “Zusanli-Shangjuxu” position of recording electrode. Thereafter, the rats were
points. The stimulation was set as a square wave pulse with a euthanized and perfused through the heart with 4% of
width of 5 ms and frequency of 20 Hz. The intensity was 1.5 paraformaldehyde. Then the rats’ brains were removed and
times of the threshold of A𝛿 fiber [14] (the average threshold fixed. After an interval of 72 hours, frozen sections of the
intensity of A𝛿 fiber reflex was 1.54 ± 0.50 mA) and the time brain were cut for H&E staining (Figure 3). Recording points
for EA was 30 s. The discharge of VPL neurons to EA was that were not located in the VPL nuclei were removed from
observed before and after CRD. the study.
M2 M1 S1Tr
RSD
DZ
S1
RSGe cg
alv
Or
IG Py S1BF
CA1 Rad
cc LMol SLu
Dbc GrDG LV
hif
PoDG S1ULp
LDDM LDVL st
3V sm
LHb cst
S2
2
PV CL
MDM MDL
MD AngT
Po
Rt CPu
PC Sc G1
CM FM
VL D1
GP
Rh SubD
SubV VM AIP
Re mt B ASe
VRe ZI EP CeL LaDL
aXi A13 ns MePD
DA MCLH CeM CeC DEn
IMG
DMD PeFLH
PLH ope STIA BLA BLP 3 2
f 1 VEn
PTe MeAD
VMHSh BMP
BMA BLV
VMHVL TuLH MeAV
a ACo 3
ArcM
ArcL 2
RChL PLCo1
Figure 3: 󳵳 refers to the location of VPL neurons.
3. Results
3.1. General Characteristics of the Responses of VPL Neurons.
A total of 126 VPL neurons that responded to mechanical
stimulations from the body surface were identified in the 26
male SD rats, by referring to the Brain Atlas of the rat [14].
Figure 3 illustrates part of the pontamine sky blue positioning
of VPL neurons. Their receptive fields were distributed at the
poster lateral of the contralateral body, tail, hips, or hind legs.
The receptive fields of most neurons were small but had clear
boundaries. The receptive fields could be activated by gentle
brushing or tapping by von Frey filaments (Figure 4).
3.2. The Influence of Different Intensities of CRD on the Dis-

charge of VPL Neurons. We isolated 54 convergent neurons
from the 126 VPL neurons that responded to inputs of
mechanical stimulation and systematically observed the dis-
charge of 9 of the 54 convergent neurons caused by different Figure 4: Distribution of the receptive fields of partial VPL neurons.
intensities of CRD stimulation. The results showed that, after
CRD stimulation ranging from 20 to 80 mmHg, the discharge
frequency of VPL neurons significantly increased in rats more
than before CRD stimulation (𝑃 < 0.01) (Figure 5). discharge of 45 convergent neurons was observed when rats
were given different intensities of CRD. Among them, 12 con-
3.3. The Influence of Different Intensities of CRD on the vergent neurons were chosen from rats receiving 20 mmHg of
Discharge Frequency of VPL Neurons Induced by EA. The nonnoxious CRD, 11 from rats receiving 40 mmHg of noxious
700 ∗∗ 800
∗ ∗
∗∗
600 700 ∗
∗
500 ∗∗ ∗∗ 600
Spikes/30 s
400 500
Spikes /30 s
300 400
200 300
100 200
0 100
20 40 60 80
CRD (mmHg) 0
20 40 60 80
Before CRD
CRD (mmHg)
After CRD
EA before CRD
Figure 5: Discharge of VPL neurons before and after CRD.
EA after CRD
Figure 6: The influence of EA on the discharge of VPL neurons

before and after CRD.
CRD, 12 from rats receiving 60 mmHg of strong nociceptive
CRD, and 10 from rats that were given 80 mmHg of strong
nociceptive CRD. 45
Equal intensities of EA were given to rats for 30 sec-
onds before and after CRD. The results showed that the
discharge frequency of VPL convergent neurons induced by
EA increased significantly after CRD more than before CRD 30
when rats were given different intensities of CRD (𝑃 < 0.05)
(%)
(Figure 6).
15
3.4. The Influence of Different Intensities of CRD on the
Discharge Number of VPL Neurons Induced by EA. After
rats were given CRD, the discharge from VPL convergence
neurons induced by EA increased over the discharge before 0
CRD: 20 mmHg–15.38% ± 8.27; 40 mmHg–25.22% + 7.80; 20 40 60 80
60 mmHg–36.28 + 8. 18; 80 mmHg–38.40 + 8.32. Differences CRD (mmHg)
were statistically significant (𝑃 < 0.05).
As the intensity of CRD stimulation increased, there also Figure 7: The influence of different intensities of CRD on VPL
was an increase in the percentage of the discharge number neuronal discharge induced by EA.
of VPL neurons from EA at acupoints. A certain dose-
effect relationship could be observed between stimulation
and response. It showed that acupoints on the body surface Results of this study showed that, within a certain inten-
were sensitized after CRD. The effect of EA on acupoints sity range, the discharge frequency of VPL convergent neu-
was enhanced. The sensitization of acupoints increased as the rons increased as the intensity of CRD stimulation increased.
intensity of visceral noxious stimulation increased (Figure 7). Since CRD had an activation effect on spinal cord neurons,
The above results showed that noxious visceral stimula- when EA was applied at acupoints after CRD, the discharge of
tion facilitated the responses of VPL neurons to inputs of EA VPL convergent neurons had a significant increase more than
stimulation from acupoints on body surface. before CRD. This confirms that noxious visceral distension
can sensitize VPL neurons making them respond more
4. Discussion strongly to inputs from EA applied to acupoints on skin
receptive fields. In other words, the neural facilitation of VPL
Our previous studies have shown that most neurons which neurons after noxious visceral stimulation led to dynamic
responded to somatic afferent inputs also responded to changes of the response from the acupoint sensitized. As
inputs from CRD or skin vibrotactile stimulation. In most the intensity of visceral noxious stimulation increased, its
cases, the response was shown as sensitization of neurons. effect on sensitization of acupoints on body surface also
Responses of more than 50% of neurons to skin vibrotactile strengthened and showed a clear dose-response relationship.
stimulation could be enhanced by CRD previously applied to Our results show that VPL neurons are involved in the
experimental animals [15]. dynamic process of acupoint sensitization.
The thalamus is the most important brain structure to depends on the group response of neurons, which includes
relay somatic and visceral afferent inputs to the cerebral cor- the interaction and feedback among nerve centers at cerebral
tex. There are three projection systems from the spinal cord cortex, thalamus, and other areas.
to the bottom part of ventral thalamus: the spinothalamic Our study showed that nociceptive stimulation of CRD
tract, the cervical spinal column, and the postsynaptic dorsal could make VPL neurons more sensitive to EA stimulation
column ascending fibers. A study by Yang et al. [16] on VPL applied at skin receptive fields. It indicates that viscera patho-
of rats showed that 94% of VPL neurons could be activated logical condition can facilitate the afferent inputs from stim-
by nonnoxious and noxious stimuli applied at peripheral ulation at the body surface. The interaction between somatic
receptive fields, whereas 6% of VPL neurons only responded and visceral inputs occurs at the lumbosacral segments of
to noxious stimulation. No VPL neurons responded only the spinal cord. The segments (L1–L3) not only integrate
to nonnoxious stimulation. Nearly 60% of VPL neurons information from the skin on lower abdomen and hind legs,
also responded to CRD, primarily with activation. VPL but also are the location of afferent neurons for “Zusanli-
neurons, therefore, are involved not only in the transmission Shangjuxu” points which were elected in our experiment and
and processing of somatic sensory inputs, but also in the dominate the lower digestive tract. Many sensitive points on
transmission and processing of visceral nociceptive inputs. body surface are distributed at relevant acupoint zones that
We observed in the rat thalamus VPL experiment that have a regulatory effect on digestive system functions. The
most neurons that responded to haptic inputs from con- phenomenon that visceral nociceptive inputs can facilitate
tralateral body also responded to CRD and skin vibration the neural responses to afferent inputs from the body surface
tactile stimuli. The responses of more than half of the neurons at corresponding spinal segments may be related with the
to skin vibrotactile stimulation could be enhanced by CRD mechanism underlying referred pain. It also provides a
conditioned stimulation applied previously. In contrast, the scientific explanation for the Chinese medical theory of “pain
responses of VPL neurons to CRD were not enhanced by skin as acupoints” and “essence of acupuncture points.”
tactile stimulation, if the order of conditioned stimulation
was reversed; that is, the skin stimulation was given before
CRD. Moreover, the effect was mainly shown as an inhibitory Conflict of Interests
effect. A possible explanation for acupoint sensitization is that The authors declare that they have no conflict of interests
repeated CRD may cause the irritability of intestinal wall, regarding the publication of this paper.
which can be viewed as one type of visceral inflammation,
and induces sensitization of afferent neurons [17]. Visceral
noxious stimulation could also significantly enhance neu- Acknowledgments
ronal responses to skin tactile stimulation. The enhancement
effect may be related with hyperalgesia caused by visceral This work was supported by the Key Project of the National
disease [18, 19]. Natural Science Foundation of China (30772830) and the 973
Many previous studies suggest that only noxious stim- Project (2011CB505201, 2012CB518503).
ulation can significantly inhibit the afferent transmission
of nociceptive inputs [18]. However, we observed that, at References
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http://dx.doi.org/10.1155/2015/671242
Review Article
Analysis and Thoughts about the Negative Results of
International Clinical Trials on Acupuncture
Wei-hong Liu, Yang Hao, Yan-jing Han, Xiao-hong Wang, Chen Li, and Wan-ning Liu
Correspondence should be addressed to Wan-ning Liu; lwn8864@163.com
Received 22 July 2014; Revised 1 December 2014; Accepted 3 February 2015
Academic Editor: Chong-Zhi Wang
Copyright © 2015 Wei-hong Liu et al. This is an open access article distributed under the Creative Commons Attribution License,
An increasing number of randomized controlled trials (RCTs) of acupuncture have proved the clinical benefits of acupuncture;
however, there are some results that have shown negative results or placebo effects. The paper carried out an in-depth analysis on
33 RCTs in the 2011 SCI database, the quality of the reports was judged according to Jadad scores, and the “Necessary Information
Included in Reporting Interventions in Clinical Trials of Acupuncture (STRICTA 2010)” was taken as the standard to analyze
the rationality of the therapeutic principle. The difference between the methodology (Jadad) scores of the two types of research
reports did not constitute statistical significance (𝑃 > 0.05). The studies with negative results or placebo effects showed the
following deficiencies with respect to intervention details: (1) incompletely rational acupoint selection; (2) inconsistent ability of
acupuncturists; (3) negligible needling response to needling; (4) acupuncture treatment frequency too low in most studies; and
(5) irrational setting of placebo control. Thus, the primary basis for the negative results or placebo effects of international clinical
trials on acupuncture is not in the quality of the methodology, but in noncompliance with the essential requirements proposed by
acupuncture theory in terms of clinical manipulation details.
1. Introduction the current study was to determine the basis for the negative
results or placebo effects in published acupuncture RCTs
As an integral part to the Chinese medical and health care from the perspective of methodology and interventions
system, acupuncture therapy is widely applied in clinical after comprehensively reading and analyzing the published
applications, effective in treatment, economical, and safe and acupuncture RCTs retrieved from the 2011 SCIE database,
thereby generally accepted by Chinese people. Since the sixth with the exception of research conducted in China.
century AD, acupuncture has successively spread to various
countries of the world, making considerable contributions
to relieving people from diseases worldwide. Along with 2. Materials and Methods
the development of evidence-based medicine, international
2.1. Search Strategy. The computer retrieval was carried out in
clinical trials on acupuncture have been increasing in num-
“Science Citation Index Expanded (SCIE)” and the retrieval
ber and raising greater controversies on whether or not
type was “(‘acupuncture’ [MeSH Terms] OR ‘acupuncture’
acupuncture is effective. While the majority of international
[All Fields] OR ‘acupuncture therapy’ [MeSH Terms] OR
clinical trial reports on acupuncture have demonstrated
(‘acupuncture’ [All Fields] AND ‘therapy’ [All Fields]) OR
that acupuncture therapy is indeed effective, some research
‘acupuncture therapy’ [All Fields] OR (‘moxibustion’ [MeSH
has shown that acupuncture therapy benefits patients, but
Terms] OR ‘moxibustion’ [All Fields] AND ‘2011/1/1’ [PDat]:
is equivalent to the placebo effect [1], and some people
consider acupuncture therapy to be ineffective [2]. Currently, ‘2011/12/31’ [PDat] AND English [lang]).”
it is widely believed that such a result is a product of the
higher-quality methodology for international randomized 2.2. Inclusion Criteria. The inclusion criteria were as follows:
controlled trials (RCTs) of acupuncture. The purpose of (1) randomized controlled acupuncture, acupressure, or
Table 1: Jadad score standard.
Score standard
Items
0 1 2
The method of randomization
Randomization Not randomized or inappropriate The study was described as
was described and it was
method of randomization randomized
appropriate
The method of double blinding
Double blinding Not blind or inappropriate The study was described as
was described and it was
method of blinding double blind
appropriate
Withdraws and drop outs A description of withdraws and
Not describing the follow-up —
dropouts
Records identified through

database searching (n = 867)
Records excluded
(n = 507)
(1) Animal experiments (n = 183)
Records after duplicates removed (2) Review (n = 26)
(n = 266) (3) First author or trial has taken
place in China (n = 198)
Records screened (n = 601)
Full-text articles assessed for

eligibility (n = 94) Full-text articles excluded, with
reasons (n = 61)
(1) Nonrandomized trial (n = 42)
(2) Intervention was not
eligible (n = 19)
Studies analyzed (n = 33)
Figure 1: The flow diagram of the research.
moxibustion trials published in 2011 from the SCIE database; 2.4. Data Collection and Analysis. Evaluation was performed
(2) patients underwent the trial regardless of age, gender, independently by two authors (Yang Hao and Wan-ning Liu).
ethnicity, or course of disease; and (3) intervention of the Relevant full articles were sorted and cross-examined. Any
observation or controlled group was based on the theory of discrepancies were discussed or further evaluated by a 3rd
meridians and collaterals and the patients were treated by author (Wei-hong Liu). Methodology was evaluated based on
acupuncture, acupressure, and/or moxibustion. the Jadad score [3]. The specific evaluation standard is shown
in Table 1.
2.3. Exclusion Criteria. The exclusion criteria were as follows: 3. Results
(1) nonrandomized trials; (2) nonclinical trials; (3) the inter-
vention did not conform to the objective of the research; (4) 3.1. Articles Included. Of the 867 articles retrieved, 33 studies
duplicated articles; and (5) the first author was from China or [2, 4–35] met the inclusion criteria for the current analysis
the trial was conducted in China. (Figure 1).
3.2. Features of the Studies. Features of the included trials are in the 17 reports were improper. For example, in item 001,
detailed in Table 2. moxibustion was adopted to treat constipation; in item 009,
which involved the treatment of postmenopausal women
3.3. Jadad Score of the Trials. According to the research suffering knee joint pain, women without medical knowledge
results, the 33 reports were classified into two types (positive performed acupressure by themselves at home; and in item
results and negative results or placebo effects). The 33 reports 005, when treating the nausea and vomiting associated with
were read and the key points were extracted. According to labor and delivery, only a wrist band that slightly stimu-
the Jadad score, the lowest methodology quality was scored lates PC 6 was used. Despite certain therapeutic function,
0, while the highest methodology quality was scored 5. The these measures are all not the most proper choice. For
clinical trial was considered low in quality if the score was example, constipation is most often treated clinically with
≤2 and was considered high in quality if the score was ≥3. acupuncture; however, for excess syndrome or heat syndrome
The Jadad scores of the research report methodologies are constipation, it is improper to use moxibustion. Similarly,
shown in Table 3, and the Jadad score comparison of the it is doubtful that the wrist band which stimulates PC 6 is
acupuncture RCT methodologies is shown in Table 4. satisfactory to achieve a therapeutic effect like acupuncture.
By adoption of SPSS 13.0, data in Table 4 was subjected to With respect to the studies with positive results, the applied
2
a 𝜒 test; the difference between the two groups was not sta- methods were effective intervention, such as filiform needles,
tistically significant (𝑃 = 1.0). The quality of the clinical trial electroacupuncture, blunt needles, or auricular acupuncture.
report methodology on acupuncture with positive results is Clearly, effective intervention is an important factor for the
similar to the clinical trial reports with negative results or results of the trial.
placebo effects, which indicates that the difference in quality
of the methodology is not the primary reason for the different 3.4.2. Acupoint Selection in Some Studies Is Not Completely
clinical research results of acupuncture. Rational. Each acupoint has its therapeutic effect. Rich expe-
riences in acupuncture have been accumulated through the
3.4. Analysis of Acupuncture RCTs Intervention Details with inheritance for thousands of years in China. For example, the
Positive Results, Negative Results, and Placebo Effects. The most effective acupoints for constipation treatment are ST25,
intervention details of the RCTs with positive results, negative ST36, ST28, ST29, and TE6, while the researcher for item
results, and placebo effects were compared. Due to the small 001 selected ST23 and ST27. Clinically, ST23 is often involved
number of RCTs with negative results or placebo effects in the treatment of gastric diseases or mental diseases, such
collected in 2011, the acupuncture RCTs from 2005 to 2010 as vexation and manic-depressive psychosis, while ST27 is
in the SCIE database were retrieved. The retrieval terms were mainly used in the treatment of hypogastrium distention
as follows: “acupuncture” [MeSH terms] OR “acupuncture” and fullness, difficult urination, hernia, spermatorrhoea,
[all fields] OR “acupuncture therapy” [MeSH terms] OR premature ejaculation, and other male diseases. The selection
“acupuncture” [all fields] AND “therapy” [all fields] OR of improper acupoints results in a low clinical curative effect.
“acupuncture therapy” [all fields] OR “moxibustion” [MeSH In item 003, the researcher only selected LI4 to treat infantile
terms] OR “moxibustion” [all fields] AND “2005/1/1” [PDat]: colic. LI4 belongs to the large intestine meridian and plays
“2010/12/31” [PDat] AND randomized controlled trial [PDat] a role in treating intestinal disease, but it is clinically known
AND English [lang]. Seven reports [36–42] with negative for its effect on head/face diseases, sweating disorders, and
results or placebo effects were analyzed together with the 10 gynecologic diseases, including menstrual disorders, vaginal
reports with negative results or placebo effects. discharge, and parturition. If GV12 and ST36 are added in the
According to the “Necessary Information Included in prescription, the effect would be significantly improved. Thus,
Reporting Interventions in Clinical Trials of Acupuncture the researcher may select a single acupoint by reducing con-
(STRICTA 2010),” the authors designed an intervention table founding factors in interventions as much as possible, but not
for the RCTs and compared the intervention details of the considering that acupuncture therapy needs the compatibility
RCTs with positive results, negative results, and placebo of acupoints for enhancement of effect and improvement
effects. The detailed information of the reports is shown in of curative effect in a practical environment. The acupoints
Tables 5 and 6. selected in the studies with positive results were correct
It is known that the clinical treatment process of and most effective according to clinical experiences. The
acupuncture involves not only the operation of acupuncture– comparison indicated that the rational selection of acupoints
moxibustion therapy, but also the rational selection of thera- is directly related to the validity of the trial.
peutic principles and methods, rational application of acu-
points, manipulation, and the correct setting of the therapy, 3.4.3. Needling Response Is Neglected in Most Studies. The
and so on. It is apparent from the analysis of items displayed famous acupuncture work, Biaoyoufu, in ancient China says,
in Tables 5 and 6 that the intervention process in the negative “Quick needling response results in quick action, otherwise
results and placebo effects was replete with defects. the late needling response causes treatment failure,” which
means that the patients’ meridian-qi circulation should be
3.4.1. Interventions of Some Trials Are Improper. To select a considered when needling to realize the needling response.
proper therapeutic method is the key to assuring a curative Of 17 reports with negative results or placebo effects, 65%
effect; however, the authors showed that some interventions (11/17) did not mention whether or not the needling response
4
Table 2: Features of the studies.

Intervention of the Intervention of the Sample Follow-up
Number Title Disease Outcome
observation group control group size phase
Heat insulation
moxa-moxibustion
“The Effectiveness of Moxibustion for the Treatment of
(place a heat
001 Functional Constipation: A Randomized, Constipation Moxibustion 26 Invalid 2 weeks
insulator under the
Sham-Controlled, Patient Blinded, Pilot Clinical Trial”
true
moxa-moxibustion)
“The efficacy of Acupressure at the Sanyinjiao Point in Both valid, the No
002 Woman health Acupressure at SP 6 Sham acupuncture 86
the Improvement of Women’s General Health” real is the better mention
Press the
nonacupoint (not
“Preliminary Research Article Electroacupuncture Is
Electroacupuncture touch the tendo No
003 Not Effective in Chronic Chronic neuralgia 16 Invalid
for 12 weeks calcaneus in any mention
Painful Neuropathies”
meridian vessel on
the back of leg)
The symptoms
“Feeding, Stooling and Sleeping Pattern in Infants with
improved, but No
004 Colic- A Randomized Controlled Trial of Minimal Infantile colic Acupuncture Blank 90
no statistical mention
Acupuncture”
difference
“The Effect of Acupuncture on Psychosocial Outcomes Women
No
005 for Women Experiencing Infertility: A Pilot experiencing Acupuncture No intervention 32 Valid
mention
Randomized Controlled Trial” infertility
“Effect of Acupuncture on Nausea and/or Vomiting Nausea and Wrist band Wrist band
No
006 during and after Cesarean Section in Comparison with vomiting when stimulative to nonstimulative to 450 Invalid
mention
Ondansetron” delivery acupoint PC 6
Conventional
“Acupuncture in Children and Adolescents with
007 Bronchial asthma Acupuncture treatment of western 93 Partial response 4 months
Bronchial Asthma: A Randomised Controlled Study”
medicine
Stick on the skin by
“True and Sham Acupuncture Produced Similar
Polycystic ovarian sham needle tubing
008 Frequency of Ovulation and Improved LH to FSH Electroacupuncture 96 Both valid 3 months
syndrome and adjust the
Ratios in Women with Polycystic Ovary Syndrome”
current to 0
“Effects of Auricular Acupuncture on Anthropometric,
Auricular Sham auricular No
009 Lipid Profile, Inflammatory and Immunologic Markers: Obesity 204 Valid
acupuncture acupuncture mention
A Randomized Controlled Trial Study”
“Acupuncture for “Frequent Attenders” with
Conventional
010 Medically Unexplained Symptoms: Neurosis Acupuncture 80 Valid 52 weeks
treatment
A Randomised Controlled Trial (CACTUS Study)”
Sham acupuncture
Someone has
“The Effects of Acupuncture on the Inner Ear Inner ear originated (the Park sham No
011 Acupuncture 63 valid in a short
Originated Tinnitus” tinnitus acupuncture mention
time
instrument)
Table 2: Continued.
“Evaluation of Wet-Cupping Therapy for Persistent
Persistent Wet-cupping +
Non-Specific Low Back Pain: A Randomised, Excise +
012 nonspecific low back excise + 32 Valid 2 weeks
Waiting-List Controlled, Open-Label, Parallel-Group analgesic-antipyretic
pain analgesic-antipyretic
Pilot Trial”
“Acupuncture in Preterm Babies during Minor Painful Without any No
013 Acupuncture Acupuncture 10 Valid
Procedures” measure mention
“Moxibustion for Cephalic Version: A Feasibility Without any No
014 Malposition Moxibustion 20 Invalid
Randomised Controlled Trial” measure mention
“Patient Education Integrated with Acupuncture for Patient education
Cancer-related No
015 Relief of Cancer-Related Fatigue Randomized integrated with Self-management 13 Valid
fatigue mention
Controlled Feasibility Study” acupuncture
Minimizing
“Effectiveness of Acupressure and Acustimulation in
driving simulation Acupressure and Sham wristband No
016 Minimizing 25 Valid
adaptation acustimulation stimulation mention
Driving Simulation Adaptation Syndrome”
syndrome
“Comparative Effects of Acupressure at Local and Distal Valid and the
Acupuncture at
Acupuncture Points on Pain Conditions and Acupuncture at local local points

017 Chronic neck pain distal points and 33 No
Autonomic Function in Females with Chronic Neck points intervention is
blank control
Pain” better
Cervical ripening
“Electroacupuncture for Cervical Ripening Prior to No
018 prior to labor Electroacupuncture Moxibustion 72 Valid
Labor Induction: A Randomized Clinical Trial” mention
induction
Valid in relieve
of the pain and
the mortality
“Effects of Acupuncture in Reducing Attrition and HIV-infected men
reduction, but
019 Mortality in HIV-Infected Men with Peripheral with peripheral Acupuncture Sham acupuncture 114 14 weeks
the severe is not
Neuropathy” neuropathy
better than
sham
acupuncture
“Acupuncture Versus Paroxetine for the Treatment of
Premature
020 Premature Ejaculation: A Randomized, Acupuncture Paroxetine 90 Valid No
ejaculation
Placebo-Controlled Clinical Trial”
“Ear Acupuncture in the Treatment of Migraine Acupuncture on
Acupuncture on 120
021 Attacks: A Randomized Trial on the Efficacy of Migraine attacks inappropriate 94 Valid
auricular point minutes
Appropriate Versus Inappropriate Acupoints” auricular point
Acupuncture care as
“Cost-Effectiveness of Acupuncture Care as an Adjunct
Osteoarthritis of the an adjunct to Education and
022 to Exercise-Based Physical Therapy for Osteoarthritis of 352 Valid 12 months
knee exercise-based excise
the Knee”
physical therapy
5
6
Table 2: Continued.
“A Randomised, Double-Blinded, Placebo-Controlled Nausea and Wrist band Wrist band
No
023 Study of Acupressure Wristbands for the Prevention of vomiting when stimulative to nonstimulative to 340 Invalid
mention
Nausea and Vomiting During Labour and Delivery” delivery acupoint PC 6
“Effect of Acupuncture on Allergen-Induced Basophil
No
024 Activation in Patients with Atopic Eczema: A Pilot Atopic eczema Acupuncture Blank 10 Valid
mention
Trial”
Sham acupuncture
(2–4 cm beside
“Acupuncture to Treat Primary Dysmenorrhea in Primary
025 Acupuncture acupoints and 92 Valid 1 year
Women: A Randomized Controlled Trial” dysmenorrhea
Streitberger sham
acupuncture)
Sham acupuncture
“Getting the Grip on Nonspecific Treatment Effects: Both better than
(2 cm beside No
026 Emesis in Patients Randomized to Acupuncture or Emesis Acupuncture 277 conventional
acupoints and park mention
Sham Compared to Patients Receiving Standard Care” therapy
sham acupuncture)
“Perioperative Acupuncture and Postoperative
Acupuncture +
Acupressure Can Prevent Postoperative Vomiting Postoperative Conventional
027 conventional 154 Valid Yes
following Paediatric Tonsillectomy or Adenoidectomy: vomiting therapy
therapy
A Pragmatic Randomised Controlled Trial”
Postmenopausal Sham acupuncture
“The Effect of Acupuncture on Postmenopausal
symptoms and (the Streitberger
028 Symptoms and Reproductive Hormones: A Sham Acupuncture 55 Valid No
reproductive sham acupuncture
Controlled Clinical Trial”
hormones instrument)
“Efficacy of Acupuncture in Preventing Atrial
029 Atrial fibrillation Acupuncture Sham acupuncture 54 Valid 12 months
Fibrillation Recurrences after Electrical Cardioversion”
Sham acupuncture
“Acupuncture for the Induction of Labour: A (the Park sham
030 Accouching Acupuncture 125 Invalid Yes
Double-Blind Randomised Controlled Study” acupuncture
instrument)
“Comparison between the Effects of Trigger Point
The trigger
Mesotherapy Versus Acupuncture Points Mesotherapy Lumbago and Lidocaine injection Lidocaine injection
031 62 point mesoderm 12 weeks
in the Treatment of Chronic Low Back Pain: A Short backache at acupoint at trigger point
has good effect
Term Randomized Controlled Trial”
High frequency of
Relaxation
“Relaxation Acupressure Reduces Persistent Cancer-related Relaxation acupuncture and No
032 43 acupressure
Cancer-Related Fatigue” fatigue acupressure low frequency of mention
valid
acupuncture
Acupuncture and Sham acupuncture
“Delayed Effect of Acupuncture Treatment in OA of the One
033 OA of the knee conventional and conventional 55 Valid
Knee: A Blinded, Randomized, Controlled Trial” month
therapy therapy
Table 3: Jadad scores of the research.
Research Withdraws Jadad

Randomization Double blinding
and drop outs score
Computer generated random table
Park et al. [4] in a 1 : 1 ratio with block size 4, and Patient blinded 2 5
using a sealed envelope
Kashefi et al. [2] Randomized Single blinded 10 3
Penza et al. [5] Randomized Patient and examiner blinded Not mentioned 2
Landgren et al. [6] Not mentioned Nurse and parents blinded 5 2
Smith et al. [7] Computer generated randomization
Statistician blinded 2 4
schedule
El-Deeb and Ahmady [8] Not mentioned Double blinded Not mentioned 3
Scheewe et al. [9] Not mentioned Not mentioned 27% 2
Pastore et al. [10] Block randomization double blinded 14 4
Hamid et al. [11] Not mentioned Not mentioned 35 1
Paterson et al. [12] Simple randomization Statistician blinded 3,1 4
Rogha et al. [13] Not mentioned Not mentioned 9 1
Kim et al. [14] Block randomization open 3 3
Ecevit et al. [15] Not mentioned Not mentioned Not mentioned 0
Do et al. [16] Computer generated randomization Not mentioned 1 2
Johnston et al. [17] Block randomization Open 1 3
Cox et al. [18] Not mentioned Not mentioned 1 0
Matsubara et al. [19] Not mentioned Not mentioned Not mentioned 0
Gribel et al. [20] Block randomization Open 0 2
Shiflett and Schwartz [21] Randomized Patient and assessor blinded 19 4
Sunay et al. [22] Simple randomization Single blinded 0 3
Allais et al. [23] Simple randomization Single blinded 1 2
Whitehurst et al. [24] Not mentioned Researcher blinded 49 0
Sinha et al. [25] Simple randomization Double blinded 11 4
Pfab et al. [26] Block randomization Researcher blinded 0 3
Smith et al. [27] Block randomization Patient and assessor blinded 2 3
Enblom et al. [28] Not mentioned Assessor and nurse blinded 32 2
Liodden et al. [29] Block randomization Double blinded 32 4
Sunay et al. [30] Not mentioned Single blinded 2 2
Lomuscio et al. [31] Patient, assessor, and statistician
Not mentioned 0 3
blinded
Modlock et al. [32] Patient, assessor, and statistician
Block randomization 19 4
blinded
di Cesare et al. [33] Block randomization Assessor blinded 2 3
Zick et al. [34] Computer generated randomization Patient blinded 8 3
Lev-Ari et al. [35] Not mentioned Patient and assessor blinded 14 2
Table 4: Jadad score comparison of the acupuncture RCT methodologies in the 2011 SCI database.
Result Total number Number of reports Proportion Number of reports Proportion

of reports scoring at 1-2 (%) scoring at 3–5 (%)
Positive 23 9 39.13% 14 60.87%
Negative/placebo 10 4 40.00% 6 60.00%
8
Table 5: Intervention of 17 research reports with negative results or placebo effects.

Needling
Number Disease Intervention Acupoint Treatment frequency Qualification of acupuncturist
response
Experience of more than five
1 Constipation Moxibustion ST 23, ST 27 3 times/week Unmentioned
years
Sham-electroacupuncture
2 Chronic neuralgia after electroacupuncture for 12 ST36, SP6, LR3, BL60 1 time/week Unmentioned Unmentioned
weeks
3 Infantile colic Acupuncture LI 4 2 times/week Unmentioned Certificated nurse practitioner
Operation by patients
4 Malposition Moxibustion BL 67 2 times/day Unmentioned
themselves who are trained
Nausea and vomiting Wrist band stimulating to
5 PC 6 1 time/week Unmentioned Unmentioned
when delivery acupoint
Acupuncturists and midwives,
6 Aids to delivery Acupuncture BL 67, LI 4, SP 6 and DU 20 2 times/day Unmentioned who often implement
acupuncture treatment
Lumbago and GB30, BL31, BL52, GV3, ā shı̀ points, 3-year training and 8-year
7 Lidocaine acupoint injection 1 time/week Unmentioned
backache GB34, GB 41, BL60, KI4, TE5 clinical experience
Selecting 6 to 10 acupoints from SP9, 67 physical therapists reaching
Physical therapy plus verum SP10, ST 34, ST35, ST36, EX-LE5, GB the lowest acupuncture level
8 Knee osteoarthritis 2 times/week Yes
acupuncture 34, ā shı̀ points, remote end: LI 4, SP 6, required by Acupuncture
LR 3, ST 44, KI 3, BL 60, and GB 41 Association.
Knee joint pain of
Routine nursing plus EX-LE4, ST35, SP 10, ST 34, ST 36, SP Operation by patients
9 postmenopause 1 time/day Unmentioned
acupressure 9, GB 34, and EX-LE2 themselves
women
Routine treatment plus Operation according to traditional Eligible acupuncturists having
10 Emergency treatment 1 time/day Unmentioned
acupuncture Chinese medical standards the experience of 6–22 years
Irritable bowel 8-16 acupoints (no specific acupoints Members approved by British
11 Acupuncture 1 time/week Yes
syndrome are mentioned) Acupuncture Association
Every day during menstrual Trained about finger force
12 Woman health Acupressure at SP 6 SP 6 Unmentioned
period practice
Two times/week within the first
Polycystic ovarian Electroacupuncture: BL23 on two Acupuncturists with the
13 Electroacupuncture four weeks, and one time/week Unmentioned
syndrome sides, BL 28, SP 6, and SP 9 experience of five years
within later four weeks
Three times/week to two
14 Emesis Acupuncture PC 6 Yes Unmentioned
times/week
2000-hour training and
Irritable bowel
15 Acupuncture CV10, ST25, LR 3, SP4, PC6, ST 37 Unmentioned Yes experience of more than four
syndrome
years
Physicians from various majors,
16 Low back pain Acupuncture Unmentioned Two times/week Yes trained about acupuncture
more than 140 hours
17 Smoking cessation Acupuncture HT 7, PC 7, HT 8, KI 3, and KI 6 Three times/week Yes Unmentioned
Table 6: Intervention of 23 research reports with positive results.
Treatment Needling Qualification of

Author Disease Intervention Acupoint
frequency response acupuncturist
Certificated
PC 6, PC 5, HT 5, HT acupuncturist,
Smith et al. [7] Infertility Acupuncture 1 time/week Yes
7 experience of more than
14 years
Nausea and
El-Deeb and
vomiting after Electroacupuncture PC 6 Single time Unmentioned Unmentioned
Ahmady [8]
cesarean
BL 13, CV 17, LU 7
and acupoint
Bronchial Experience of many
Scheewe et al. [9] Acupuncture selection according to 3 times/week Yes
asthma years
syndrome
differentiation
Auricular HT 7, CO4, CO1,
Hamid et al. [11] Obesity 2 times/day Unmentioned Unmentioned
acupuncture HX1, CO17
According to Member of British
Acupuncture and
Paterson et al. [12] Psychoneurosis Not mentioned syndrome Yes Medical Acupuncture
moxibustion
differentiation Society
Endogenous TE 17, GB 2, SI 19, and
Rogha et al. [13] Acupuncture 3 times/day Unmentioned Unmentioned
tinnitus TE 21
3-year training and
Kim et al. [14] Low back pain Cupping BL 23, BL 24, BL 25 3 times/week Unmentioned 6-year clinical
experience
Ecevit et al. [15] Analgesia Acupuncture EX-HN3 1 time/week Unmentioned Qualified acupuncturist
Phd of TCM in the US,
Kim et al. [14] Cancer fatigue Acupuncture LI 4, SP 6, ST 36, KI 3 Not mentioned Yes with 20 years of clinical
experience
Auricular
Cox et al. [18] Motion sickness TF4 Single time Unmentioned Unmentioned
acupuncture
Matsubara et al. GB 21, SI 14, SI 15, LI
Neck pain Acupressure Single time Unmentioned Unmentioned
[19] 4, LI 10, LI 11
Cervical LI 4, ST 36, LR 3, SP With 20 years of clinical
Gribel et al. [20] Electroacupuncture 3 times/day Unmentioned
dilatation 6, BL 23, BL 32 experience
SP 6, SP 7, SP 9, and
Peripheral acupoint selection
Shiflett and Has received
neuropathy in Acupuncture according to 2 times/week Unmentioned
Schwartz [21] standardized training
ADIS patients syndrome
differentiation
Certificated and
Premature LI 4, ST 36, KI 3, LR
Sunay et al. [22] Acupuncture 2 times/week Unmentioned experienced
ejaculation 3, EX-HN3, CV 3
acupuncturist
Experienced
Allais et al. [23] Migraine Blunt needle AT4 Single time With
acupuncturist
Conventional Member of British
Whitehurst et al. Knee
Acupuncture acupoints used for Not mentioned Yes Medical Acupuncture
[24] osteoarthritis
knee osteoarthritis Society, physiotherapist
LI 11, LI 4, ST 36, SP Experienced
Pfab et al. [26] Eczema Acupuncture 2 times/week Unmentioned
10 acupuncturist
Certificated
SP 4, CV 3, ST 29, SP
Smith et al. [27] Dysmenorrhea Acupuncture 3 times/week Yes acupuncturist of
6, BL 32, SP 8
CMASA
Liodden et al. Postoperative Acupuncture + Experienced
PC 6 For 24 hours Unmentioned
[29] nause acupressure acupuncturist
Perimenopausal ST 36, LI 4, KI 3, LR Certificated
Sunay et al. [30] Acupuncture 2 times/week Yes
syndrome 3, EX-HN3, CV 3 acupuncturist of 6 years
Table 6: Continued.
Treatment Needling Qualification of
Author Disease Intervention Acupoint
frequency response acupuncturist
Lomuscio et al. Atrial
Acupuncture PC 6, HT 7, BL 15 1 time/week Unmentioned Trained acupuncturist
[31] fibrillation
ST 36, SP 6, KI 3, LI 4,
Certificated
CV 6 GV 20, 2 times a day, 3
Zick et al. [34] Cancer fatigue Acupressure Yes acupuncturist with B.S.
EX-HN3, HT 7, LR 3, times/week
degree
SP 6
GB 34, SP 5, ST 35,
Knee
Lev-Ari et al. [35] Acupuncture EX-LE5, LI 4, local 2 times/week Unmentioned Unmentioned
osteoarthritis
points, ST 43, ST 34
was achieved. This is a problem that should be addressed 3.4.5. The Acupuncture Treatment Frequency Is Too Low in
by the research design and execution staff. The physicians Most Studies. Among the 17 studies listed in Table 5, eight
of acupuncture and moxibustion in China mostly have such had a treatment frequency of 1-2 times/week (002, 003, 005,
experiences that immediately after needle insertion they 007, 008, 011, 013, and 016), accounting for 47% of the studies;
must observe the patient’s response, earnestly feel the sense 53% of the studies had a treatment frequency ≥3 times/week
beneath the needle tip, and repeatedly operate the needle (001, 014, and 017). Among the studies with positive results,
body so that the endurable feelings of sourness, numbness, eight had a treatment frequency of 1-2 times/week, account-
swelling, heaviness, and pain can be felt by the patients. ing for 35%; 65% of studies with positive results had a treat-
Meanwhile, the operator also feels heaviness and tightening ment frequency ≥3 times/week. Indeed, the studies with pos-
beneath the needle tip, which is called the needling response. itive results had a significantly higher treatment frequency.
If such a feeling is generated, a good curative effect can be According to Cai and Ma [44], the influence of acupuncture
realized, whereas if it is not, the effect is slow or not apparent. at BL23 on urinary function peaks after the acupuncture
As an intervention in the observation group, no needling is implemented for 1 hour and then slowly declines and
response suggests no real or ineffective stimulation to the recovers to the original level, with the effect lasting 2–6 hours.
acupoint. In this way, the curative effect will be reduced This finding is consistent with the metabolic principles
greatly and a negative result may be more likely to occur. It in the human body. The curative effect of acupuncture is
is noteworthy that 65% (15/23) of the studies with positive determined by the duration of the acupuncture effect remain-
results did not mention whether or not the needling response ing in the human body and the accumulation of multiple
was achieved. As a complicated intervention, the effectiveness therapeutic effects. Therefore, the best treatment frequency of
of acupuncture is influenced by multiple factors. acupuncture is 1-2 times per day. In the event of one treatment
per 2 days or an extended interval of time, it takes more
3.4.4. The Requirements of the Acupuncturist Is Neglected in time to accumulate the acupuncture effect, leading to a slower
Many Studies. As shown in Table 5, acupuncturists involved onset of effect. Moreover, different diseases require different
in the clinical trials had inconsistent qualifications. The treatment frequencies; for chronic diseases and permanent
proportion of excellent acupuncturists in the studies with symptoms, the treatment frequency should be higher, and
negative results or placebo effects was 65% (11/17). Some of for chronic neuralgia (001), irritable bowel syndrome (011),
the acupuncturists work part-time and are actually nurse and smoking cessation (017), it is evident that a good effect is
practitioners (003), some have achieved the lowest require- difficult to realize if the frequency is one time per week.
ments (008), some are midwives without acupuncturist In addition to all the factors above, based on the research
qualifications (006), and some ask the patients to operate on demonstrating a clinical curative effect, the diseases which
themselves at home (004, 009, and 012). Because of low-level are best to be treated are selected. For example, for smoking
acupuncturists and such simple treatments, it is really difficult cessation, a worldwide problem which is difficult to eradicate,
to fully realize the curative effect of acupuncture. The propor- if acupuncture is adopted at a frequency of one time per week,
tion of excellent acupuncturists in the studies with positive the effect is weak.
results was 74% (17/23), which was significantly higher
than the studies with negative results and placebo effects. 3.5. Reflections on Placebo Acupuncture Settings. The 17
Acupuncture is a therapeutic method that has high technical studies with negative results or placebo effects are generated
skill requirements. He [43] concluded that the proficiency in comparison with other therapeutic measures. At the same
and level of clinical acupuncture skill constitute decisive time, the suitability of the control settings is also worthy
factors of a clinical curative effect, as well as the advantages of of further analysis. The authors have analyzed the control
famous veteran physicians of traditional Chinese medicine. setting list (Table 7) in these research reports and divided
Inexperienced or unqualified acupuncturists undoubtedly the control methods into the following three types: (1) no
lower the effectiveness and safety of acupuncture treatment, penetration into the skin (the Park sham needle) or heat
especially when patients are asked to treat themselves. insulation acupuncture; (2) slight penetration into the skin
Table 7: Control design in research report with negative or placebo result.
Number Intervention of observation

Disease Intervention of control group
group
1 Heat insulation moxa-moxibustion (place a heat insulator
Constipation Moxibustion
under the true moxa-moxibustion)
2 Electroacupuncture for 12
Chronic neuralgia Sham electroacupuncture (around the acupoint)
weeks
3 Infantile colic Acupuncture Without any measure
4 Malposition Moxibustion Without any measure
5 Nausea and vomiting when Wrist band stimulative to
Wrist band nonstimulative to PC 6
delivery acupoint
6 Accouching Acupuncture Sham acupuncture (the Park sham acupuncture instrument)
7 Lidocaine injection at
Lumbago and backache Lidocaine injection at trigger point
acupoint
8 Physiotherapy and verum Physiotherapy and placebo acupuncture, or just
Knee osteoarthritis
acupuncture physiotherapy
9 Knee joint pain of
Usual care and acupressure Usual care
postmenopausal women
10 Conventional treatment
Emergency treatment Conventional therapy
and acupuncture
11 Sham acupuncture (unrelated to meridian points, without
Irritable bowel syndrome Acupuncture
needling response)
12 Press the nonacupoint (not touch the tendo calcaneus in any
Woman health Acupressure at SP 6
meridian vessel on the back of leg)
13 Polycystic ovarian Stick on the skin by sham needle tubing, adjust the current to
Electroacupuncture
syndrome 0
14 Acupuncture and Antinausea drug or sham acupuncture (two inches close to
Emesis
moxibustion the acupoint by Park sham acupuncture instrument)
15 Irritable bowel syndrome Acupuncture Park sham needle, the nonacupoint close to acupoints
16 Low back pain Acupuncture Nonacupoint shallow penetration (1–3 mm)
17 Smoking cessation Acupuncture Nonacupoint shallow penetration (1–3 mm)
or press; and (3) stimulation of the nonacupoint parts. These group is false-negative. The Park sham acupuncture method
three points will be analyzed one-by-one as follows. is similar to a pressing method. The acupressure is referred to
as the “indicator” in acupuncture theory and exclusively used
3.5.1. No Penetration into the Skin as a Control. Park sham for infants, people afraid of acupuncture, nervous patients,
acupuncture instruments were used in items 006, 013, 014, or when the needle is lacking; acupressure is also a simple
and 015, which is the control that did not penetrate into method with a treatment effect.
the skin. The instrument incorporates a round and blunt
needle head which can be retracted into the needle handle 3.5.2. Slight Penetration into the Skin as a Control. The 016
and does not penetrate into the skin when the needle is and 017 studies carried out the control using the shallow
touching the skin. The outer surface of the needle is fixed stimulation method; however, in clinical acupuncture and
by double-faced adhesive tape and equipped with a small moxibustion, shallow acupuncture itself is an effective ther-
pipe to prevent the patient from seeing the truth. Park et al. apeutic method. The Miraculous Pivot has recorded that the
[45] reported that the needle head would inevitably stimulate light stimulation just stimulates the skin, while the semipen-
the skin and have a vivid effect on the skin, which will etration involves the skin, but not the muscle. The A-B Classic
result in a physiologic effect. The Park sham acupuncture of Acu-moxibustion has clearly described that 14 acupoints
changes the method and tools of stimulation; thus the control can be penetrated by one fen (approximately 3.3 mm) and
method can also generate some therapeutic effect, but the 20 acupoints can be penetrated by 2 fen (approximately
researcher considers it as the control measure that cannot 6.6 mm) [46]. Another study [47] indicates that 42 patients
generate an effect or only shows a placebo effect. Therefore, with wrist myofascial pain were randomly distributed to the
when the measures of the observation group indicate the deep acupuncture group and the shallow acupuncture group
same curative effect as that of the control group, the measure with the same acupoints, and the acupuncture depth for the
of the observation group is considered to be invalid or have deep acupuncture group was 1.5 cm compared to 2 mm for the
placebo effect only. The measure of the control group has shallow acupuncture group. The McGill pain questionnaire
some therapeutic effect, so the result of the observation was used as the evaluating indicator. The scores for the two
groups before the treatment were 35.4 ± 14.53 and 34.75 ± such as incomplete rational acupoint selection, inconsistent
11.43, respectively, compared to 14.54 ± 10.88 and 22.25 ± ability of acupuncturists, negligence of the needling response
16.08 after the treatment. The results indicate that the two to needling, low frequency of the acupuncture treatment, and
groups can relieve the pain, and the curative effect was not irrational setting of placebo control. Those directly weaken
statistically different. Therefore, with respect to pain-related the positive property of the results in the observation group,
disease and disease suitable for shallow acupuncture, shallow and the setting of the placebo acupuncture control is opposite
acupuncture is unsuitable for the acupuncture control with- to the theory of acupuncture. The placebo acupuncture
out a curative effect because it will result in false-negative method has certain therapeutic effects instead of purely a
properties of the observation group. placebo effect, thereby causing the false-negative property
of the results in the observation group. It was shown that
3.5.3. Nonacupoint and Nonmeridian Acupoint as Controls. the sham acupuncture (placebo acupuncture) in the current
The researchers performing seven studies (002, 011, 012, 014, acupuncture RCTs and the placebo control method was not
015, 016, and 017) avoid or depart the known acupoints reached by consensus. The Society of Acupuncture and Mox-
and meridians as the placebo control. The question involves ibustion gradually found that the clinical trials under ideal
how many acupoints the people have in their body. The conditions are not suitable for acupuncture and moxibustion.
WHO has approved that there are 361 meridian points Seeking the clinical research methods in the practical world,
and 48 extraordinary points; however, >2200 extraordinary practical clinical research may be able to break the limit of
acupoints have been collected [48] with formal names and the placebo acupuncture control and find the advantage of
main functions. Owing to all types of unfixed a shı̀ points, it acupuncture therapy.
is easy to avoid the meridians, but hard to avoid the acupoints We can see that the current clinical research for acupunc-
when designing the nonacupoint and nonmeridian controls. ture and moxibustion still reflects many methodologic prob-
The parts avoiding the familiar meridians and acupoints are lems and is not mature in terms of theory and practices.
just defined as the nonmeridian and nonacupoint parts [49]. It is necessary to establish a clinical research method for
Furthermore, the area of the acupoints has not been mea- acupuncture and moxibustion to meet the requirements of
sured until now, and the distance between the meridian or the acupoint theory, practice features, and clinical trials so
acupoint and the nonmeridian and nonacupoint part has not that the clinical trial results for acupuncture and moxibustion
been determined. Therefore, the control with nonmeridian are scientific, comply with medical ethics, completely meet
and nonacupoint parts is highly possible to apply the “point” the treatment effect advantages of acupuncture, and promote
with a therapeutic effect as the control, and the result of the acupuncture to mainstream medicine.
observation group has a high possibility of a false-negative. The limitations of the research are as follows: (1) the
In summary, all of the three above-mentioned con- research report is of limited duration, thus this paper
trol methods showed a therapeutic effect; however, the inevitably suggests selection bias; (2) a common phe-
researchers only think the therapeutic effect was from the nomenon exists in the sector that the probability of publish-
placebo control and when the therapeutic effect of the ing of a negative article is lower than for a positive article,
observation group is similar to that of the control group, the which will cause bias to the research conclusion; and (3) the
conclusion is incorrect that the observation group therapy Jadad scale is used to evaluate the methodologic quality of the
had no effect or was equal to the placebo. The other reason article. The greatest strength of the scale is directly evaluating
for the researcher to design the placebo control like this is the verified test features related to the bias in the test effect
possibly related to the “blind.” In view of the particularity of evaluation, which is simple and clear; however, the Jadad
acupuncture, it is impossible to identify the placebo therapy score will be too general and arbitrary if most of the research
meeting the blind requirement and being similar to acupunc- is not defined, whether or not they are random or double
ture. Many experts [50–52] have written articles to discuss the blind.
methods of setting the control group in acupuncture RCTs;
however, Liu [53] suggests using modern medical methods Conflict of Interests
as the standard control and aiming at the most effective and
most advanced method in mainstream medicine to directly The authors declare that there is no conflict of interests
discover the advantage or disadvantage of acupuncture and regarding the publication of this paper.
give full attention to the medical development of acupunc-
ture. Authors’ Contribution
4. Discussion Wei-hong Liu conceived and designed the work; Yang Hao,
Wan-ning Liu, Yan-jing Han, Xiao-hong Wang, and Chen Li
By analyzing acupuncture RCTs in the SCI database, it is performed the work; Wei-hong Liu and Yang Hao wrote the
discovered that the methodologic quality of research with paper.
positive results is not different from that of research with
negative results or placebo effects. The methodologic quality Acknowledgments
is not the primary reason contributing to the difference in
research results; however, each study with negative results or The authors would like to give special thanks to Haoning
placebo effects has disadvantages on the intervention side, Ma for his help in data retrieval. This work was funded by
Institute of Acupuncture and Moxibustion, China Academy [14] J.-I. Kim, T.-H. Kim, M. S. Lee et al., “Evaluation of wet-cupping
of Chinese Medical Sciences, Grant no. ZZ07006, URL: therapy for persistent non-specific low back pain: a randomised,
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preparation of the paper. [15] A. Ecevit, D. A. Ince, A. Tarcan, M. T. Cabioglu, and A. Kurt,
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http://dx.doi.org/10.1155/2015/320701
Research Article
Efficacy of Acupuncture in Children with Nocturnal
Enuresis: A Systematic Review and Meta-Analysis of
Randomized Controlled Trials
Zheng-tao Lv,1 Wen Song,2 Jing Wu,3 Jun Yang,2 Tao Wang,2 Cai-hua Wu,4 Fang Gao,4
Xiao-cui Yuan,4 Ji-hong Liu,2 and Man Li4
1
Department of Orthopedics, Tongji Hospital, Tongji Medical College, Huazhong University of Science and Technology,
Wuhan 430030, China
2
Department of Urology, Tongji Hospital, Tongji Medical College, Huazhong University of Science and Technology,
Wuhan 430030, China
3
Key Laboratory of Environment and Health, Ministry of Education and Department of Epidemiology and Biostatistics,
School of Public Health, Tongji Medical College, Huazhong University of Science and Technology, Wuhan 430030, China
4
Department of Neurobiology, School of Basic Medicine, Tongji Medical College of Huazhong University of Science and Technology,
Wuhan 430030, China
Correspondence should be addressed to Man Li; liman7322@hotmail.com
Received 27 October 2014; Revised 30 January 2015; Accepted 31 January 2015
Copyright © 2015 Zheng-tao Lv et al. This is an open access article distributed under the Creative Commons Attribution License,
Background. Nocturnal enuresis (NE) is recognized as a widespread health problem in young children and adolescents. Clinical
researches about acupuncture therapy for nocturnal enuresis are increasing, while systematic reviews assessing the efficacy of
acupuncture therapy are still lacking. Objective. This study aims to assess the effectiveness of acupuncture therapy for nocturnal
enuresis. Materials and Methods. A comprehensive literature search of 8 databases was performed up to June 2014; randomized
controlled trials which compared acupuncture therapy and placebo treatment or pharmacological therapy were identified. A meta-
analysis was conducted. Results. This review included 21 RCTs and a total of 1590 subjects. The overall methodological qualities
were low. The results of meta-analysis showed that acupuncture therapy was more effective for clinical efficacy when compared
with placebo or pharmacological treatment. Adverse events associated with acupuncture therapy were not documented. Conclusion.
Based on the findings of this study, we cautiously suggest that acupuncture therapy could improve the clinical efficacy. However, the
beneficial effect of acupuncture might be overstated due to low methodological qualities. Rigorous high quality RCTs are urgently
needed.
1. Introduction a higher risk for psychosocial comorbidity and loss of self-

esteem. Such feelings of humiliation, guilt, and shame are
Nocturnal enuresis (NE) is a worldwide health problem also a reasonable source of heartache to the children and
frequently encountered in childhood and is defined as an their parents. The etiology and underlying physiological
involuntary voiding of urine during sleep with a frequency of mechanisms of nocturnal enuresis are multifactorial; three
at least twice a week in children, in the absence of congenital commonly proposed mechanisms to bedwetting include
or acquired defects of the central nervous system [1]. It excessive nocturnal urine production, bladder overactivity,
includes monosymptomatic nocturnal enuresis (MNE) with and a failure to awaken in response to bladder sensations [3].
no daytime urinary symptoms and nonmonosymptomatic Current first-line nocturnal enuresis therapies center on
nocturnal enuresis (NMNE) that is accompanied by daytime the aforementioned mechanisms; generally accepted treat-
urinary symptoms. Nocturnal enuresis affects 5%–10% of ments are oral pharmacological therapies including desmo-
younger school-age children [2]. Enuretic children have pressin, tricyclics, or oxybutynin and behavioral therapies
[4]. Desmopressin has been widely used for several decades, acupuncture therapy (including needles, moxibustion, acu-
and its reliable therapeutic effect has been proven to one- pressure, electroacupuncture, and acupoint injection, among
third of the unselected enuretic children. But the clinical other techniques), either alone or in conjunction with
effect cannot be maintained once the medication is stopped another kind of acupuncture therapy, without differentiating
and the side effects associated with drugs may cause the different acupuncture therapy techniques, acupoints selec-
patients to be reluctant to use them for long periods. The tion, or needle materials; (4) control group interventions:
preferred behavioral treatment is bed alarm, which needs control interventions included placebo acupuncture or oral
to be continuous and brings the enuretic children different pharmacological treatment; (5) outcome measurements: the
degrees of sleep disorders at the same time [5]. outcome measurement had to include overall clinical efficacy,
Complementary and alternative medicine (CAM) is number of wet nights per week, or maximum voided volume.
widely advocated to face the increasing demand for non-
pharmacological approaches. As a mainstream CAM therapy, 2.3. Exclusion Criteria. Exclusion criteria included the fol-
acupuncture treatment based on TCM theory has been lowing: (1) articles regarding animal experiments, review
commonly used to treat nocturnal enuresis in Chinese articles, case reports, or expert experience reports; (2)
cultures. Compared to conventional care, its safety and cost nonrandomized studies; (3) studies that compared different
effectiveness assure the maintenance of patients’ compliance. acupuncture modalities or acupoints selection; (4) exper-
Unfortunately, there is little published information to warrant imental groups that accepted complex therapy, while the
acupuncture therapy as standard treatment of nocturnal contributing factor could not be distinguished; (5) studies
enuresis. The aim of this review is to evaluate the efficacy of that were duplicates for retrieving or publishing.
acupuncture therapy in the treatment of nocturnal enuresis
when compared with placebo acupuncture or oral pharma-
2.4. Data Extraction. Two reviewers (Zheng-tao Lv and
cological treatment based on randomized controlled trials
Wen Song) reviewed each article independently and were
(RCTs).
blinded to the findings of the other reviewer. In accordance
with the predetermined inclusion criteria, two reviewers
2. Material and Methods independently performed a rigorous screening to identify
2.1. Literature Search Strategy. A comprehensive litera- qualified articles, and they extracted data independently from
ture search of the Cochrane Central Register of Con- these articles using a standardized collection form, which
trolled Trials (CENTRAL), Cochrane Database of Sys- includes first author, year of the study, sample size, nation
tematic Review (CDSR), EMBASE, ISI Web of Science, or region, baseline characteristics, methodological features
and PubMed was conducted. We also searched Chinese of the studies, quality of trial design, interventions, main
databases, including China Knowledge Resource Integrated outcome assessments, follow-up time, and withdrawal. If
database (CNKI), WanFang Data, VIP, and Chinese Biomed- the required information was not available in the included
ical (CBM) Literature database. In addition, we searched studies, we contacted the original authors by email. Any
databases that contained registered trials, such as Clinical- discrepancies between reviewers were resolved through dis-
Trials.gov (http://www.clinicaltrials.gov). All databases were cussion until a consensus was reached. The third review
searched from their inception dates up to June 2014; lan- author (Man Li) was consulted if a consensus could not be
guages were restricted to Chinese and English. The following reached.
medical subject headings or key words were used for English
databases: enuresis, nocturnal enuresis, nighttime urinary 2.5. Quality of the Studies. The methodological quality of
incontinence, bedwetting, acupuncture, electroacupuncture, the included trials was evaluated using the Jadad quality
auricular acupuncture, ear acupuncture, scalp acupuncture, scale [27]: (1) randomization (the study was described as
acupoint, moxibustion, acupressure, and acustimulation. For randomized), (2) double blinding (participant masking and
Chinese databases we used free text terms as “Zhen” or “Jiu” researcher masking), (3) reporting of the number of dropouts
or “Xue Wei” and “Yi Niao.” In addition, the bibliographies and reasons for withdrawal, (4) allocation concealment, and
of relevant systematic reviews and clinical guidelines were (5) generation of random numbers (by using computer,
manually searched. We also searched the gray literature random numbers table, shuffled cards, or tossed coins). RCTs
that included dissertations, letters, government documents, scored 1 point for each area addressed in the study design for
research reports, conference proceedings, and abstracts when a possible score between 0 and 5 (highest level of quality). The
available. The reference section for each study was also quality of all included studies was assessed by two authors
searched. (Zheng-tao Lv and Wen Song) and the articles were classified
as high quality if their Jadad score ≥4 and low quality if their
2.2. Inclusion Criteria. Inclusion criteria are as follows: (1) Jadad score ≤3. Disagreements regarding methodological
research subjects: the enrolled patients had to be diagnosed quality were resolved with discussion between reviewers.
with NE and no restrictions on race, age, or sex were
imposed; (2) study design: the included studies were required 2.6. Data Synthesis and Analysis. The meta-analysis and sta-
to be RCTs in Chinese or English aiming to assess the tistical analysis were performed by using RevMan 5.1 analyses
efficacy of acupuncture therapy for NE; (3) experimental software of the Cochrane Collaboration. We extrapolated the
group interventions: experimental group mainly received odds ratio (OR) and the associated 95% confidence interval
PubMed = 73 CBM = 942

Web of Science = 108 CNKI = 880
Identification
EMBASE = 136 WangFang = 633
Cochrane library = 29 VIP = 779
Records after duplicates removed

(n = 1936)
Screening
Records screened Records excluded

(n = 1644) (n = 1583)
Full-text articles assessed Full-text articles excluded,

for eligibility with reasons (n = 40)
Eligibility
(n = 61) Non-RCT = 17
Duplicates = 6
Unsuitable intervention = 7
Complex therapy = 6
Studies included in Unsuitable outcome = 4
quantitative synthesis
(n = 21)
Included
Studies included in
quantitative synthesis
meta-analysis
(n = 19)
Figure 1: Flowchart of the literature search and study selection.
(CI) for treatment effect. The chi-squared test and the Higgins two different kinds of acupuncture therapy; 4 studies did
𝐼2 test were used to assess the heterogeneity of the data [28]. not report the suitable outcome. Finally, 21 studies meet our
We pooled data across studies using random effect models if inclusion criteria [6–26]. Because only two of these studies
statistical heterogeneity exists; otherwise, a fixed effect model [10, 17] compared acupuncture with placebo treatment (e.g.,
will be used. Publication bias was explored via a funnel-plot without active laser light but with or without skin contact),
analysis. Begg’s test and Egger’s test were conducted when the we just used them for systematic review (Figure 1).
number of included studies is equal to or greater than 5 (Stata
Software, version 12.0). In case of heterogeneity, subgroup 3.2. The Characteristics and Methodological Quality of the
analysis was conducted. Included Trials. The characteristics of the 21 trials are sum-
marized in Table 1. These studies were published between
3. Results 2001 and 2014. Sixteen studies were published in Chinese
and five studies in English. The 21 RCTs included a total of
3.1. Literature Search Results. An initial search of RCTs 1590 patients with nocturnal enuresis: 826 patients in the
yielded 3580 potential literature citations, including 346 acupuncture group (experimental group) and 731 patients
English studies and 3134 Chinese studies, and 1936 duplicated in the control group. Age of the patients ranges from 3
articles were deleted. After screening titles and abstracts, 61 to 21 years. Nineteen studies used 2-parallel-arm group
potentially relevant studies were selected and retrieved for a designs [6, 8–12, 14–26] and two used a 3-parallel-arm
full-text assessment. Of the remaining 61 studies, 17 studies group design [7, 13]. The experimental group mainly received
were excluded because they were not RCTs; 6 articles were acupuncture therapy (including needles, moxibustion, acu-
duplicates; 7 studies took unsuitable intervention; 6 studies pressure, electroacupuncture, and acupoint injection, among
accept the complex therapy, for example, combination of other techniques). Among the 21 studies, western medicine
Table 1: Characteristics and methodological quality of included studies.
Sample size Nation/region Age (mean or Baseline EC approval Jadad

Study Study design
(n1/n2) range) score
Dong et al., 2012 China E: 8.61 (5∼12) years Adequate Not reported
RCT, parallel 2 arms 120 (60/60) 1
[6] C: 8.57 (5∼13) years
Hong et al., 2011 [7] RCT, parallel 3 arms 99 (33/33/33) China 5∼13 years Adequate Not reported 2
China E: 5∼12 years Adequate Not reported
Hui et al., 2006 [8] RCT, parallel 2 arms 67 (35/32) 2
C: 6∼11 years
Liu, 2007 [9] RCT, parallel 2 arms 60 (30/30) China 5∼12 years Not reported Not reported 1
RCT, parallel 2 arms
Prospective,
Karaman et al., Turkey E: 8.5 ± 3.2 years Adequate Yes
randomized, 83 (57/26) 3
2011 [10] C: 8.9 ± 3.3 years
placebo controlled,
single-blind study
Ding et al., 2007 China Adequate Not reported
RCT, parallel 2 arms 80 (42/38) 3∼13 years 1
[11]
Tong and Zhan, Guinea-Bissau Adequate Not reported
2009 [12]
Moursy et al., 2014 Egypt 15.7 years (range Adequate Yes
RCT, parallel 3 arms 186 (62/62/62) 3
[13] 10–21 years)
Tian and Zhong, China E: 7.58 ± 2.16 years Adequate Not reported
2008 [14] C: 8.26 ± 2.67 years
Ling and Chen, China E: 9.2 (5∼16) years Adequate Not reported
2011 [15] C: 9.1 (5∼15) years
Radmayr et al., Austria E: 8.6 (5∼16) years Adequate Yes
2001 [16] C: 8.0 (5∼14) years
RCT, parallel 2 arms
Prospective,
Radvanska et al., single-blind, Slovakia E: 8.7 ± 1.4 years Adequate Yes
29 (16/13) 4
2011 [17] randomized, C: 8.6 ± 1.3 years
placebo controlled
design
Yang et al., 2012 China Adequate Not reported
[18]
China E: 8.5 ± 0.1 years Adequate Not reported
Luo, 2010 [19] RCT, parallel 2 arms 40 (20/20) 1
C: 8.4 ± 0.2 years
Tang et al., 2012 China E: 5∼11 years Adequate Not reported
[20] C: 5∼12 years
Qiu, 2008 [21] RCT, parallel 2 arms 56 (31/25) China 3∼16 years Not reported Not reported 1
Zhu et al., 2003 China Adequate Not reported
[22]
Zhang, 2010 [23] RCT, parallel 2 arms 80 (40/40) China 3∼18 years Adequate Not reported 1
Chen and Gu, 2003 China Adequate Not reported
[24]
Yuksek et al., 2003 Turkey E: 7.67 ± 2.34 years Adequate No
[25] C: 7.41 ± 2.67 years
Hong and Zhang, China Adequate Not reported
2009 [26]
therapy (e.g., desmopressin, Meclofenoxate) was used as the The mean Jadad score of these 21 studies was 1.7, ranging
intervention method for the control group in 9 studies [6, 8, from 1 to 4 points (Table 1). Only 1 of 21 RCTs met the Jadad
9, 11, 13, 16, 22, 23, 25]; traditional Chinese medicine (TCD) criteria for high quality [17]. All of the studies included sug-
was used in 10 studies [7, 12, 14, 15, 18–21, 24, 26]; and placebo gested randomization, and 9 studies reported the method of
treatment or sham-acupuncture was used in 2 studies [10, 17]. random sequences generation [7, 8, 12–17, 20]. In that study,
The main outcome indicators reported in the included studies it was not feasible to blind the participant or the therapist.
were cure rate, improvement rate, and mean weekly number The outcome assessor was blinded in only two studies [10, 17];
of wet nights; Two studies reported maximum voided volume we considered that the outcomes and their measurements
(MVV) as outcome indicators [13, 17] (Table 2). are likely to be influenced by lack of blinding. Four studies
Table 2: Interventions and outcomes of included studies.
Duration of Follow-up Experimental Cure rate of Cure rate of
Study Control treatment Outcome measurement
treatment after treatment treatment intervention group control group
Acupoint injection
Dong et al., 2012 Western medicine: Cure rate, improvement rate, follow-up at 1
5 weeks 6 months with scraping therapy 46/60 (76.67%) 36/60 (60%)
[6] Meclofenoxate (𝑛 = 60) and 6 months
(𝑛 = 60)
Moxibustion (𝑛 = 33) Chinese patent 20/33 (60.6%)
Hong et al., 2011 [7] 1 month Not reported 8/33 (24.24%) Cure rate, improvement rate
Acupuncture (𝑛 = 33) medicine (𝑛 = 33) 19/33 (57.6%)
Western medicine:
Heat-producing Cure rate, total effective rate, follow-up at 1
Hui et al., 2006 [8] 1 month 1 year imipramine 20/35 (57.2%) 14/32 (43.8%)
needling (𝑛 = 35) month
hydrochloride (𝑛 = 32)
Enuresis patch Western medicine:
Liu, 2007 [9] 3 weeks Not reported 18/30 (60%) 9/30 (30%) Cure rate, improvement rate
(𝑛 = 30) Meclofenoxate (𝑛 = 30)
Complete improvement rate, partial
Placebo therapy: with a improvement rate, mean number of weekly

Karaman et al., Laser acupuncture
4 weeks 6 months nonlaser light source 31/57 (54.4%) 3/26 (11.5%) bedwetting episodes: the children were
2011 [10] (𝑛 = 57)
(𝑛 = 26) reevaluated 15, 30, 90, and 180 days after
treatment
Ding et al., 2007 Enuresis patch Western medicine:
1 month 3 months 25/42 (59.5%) 13/38 (34.2%) Cure rate, improvement rate
[11] (𝑛 = 42) Meclofenoxate (𝑛 = 38)
Tong and Zhan, Suspended Chinese patent
1 month Not reported 17/30 (56.7%) 10/30 (33.3%) Cure rate, improvement rate
2009 [12] moxibustion (𝑛 = 30) medicine (𝑛 = 30)
Western medicine: Cure rate, improvement rate, mean weekly
desmopressin (𝑛 = 62) number of wet nights, MVV (maximum
Moursy et al., 2014 Laser acupuncture 35/62 (56.5%)
3 months 6 months Combination therapy: 33 /62 (53%) voided volume): the patients were evaluated
[13] (𝑛 = 62) 46/82 (74%)
acupuncture + once every 2 weeks for 3 months and once
desmopressin (𝑛 = 62) every 4 weeks for 6 months
Tian and Zhong, Acupuncture Chinese patent
2 weeks Not reported 61/116 (52.59%) 47/112 (41.96%) Cure rate, improvement rate
2008 [14] (𝑛 = 116) medicine (𝑛 = 112)
Ling and Chen, Acupoint injection Chinese patent
1 month Not reported 18/30 (60%) 15/30 (50%) Cure rate, improvement rate
2011 [15] (𝑛 = 30) medicine (𝑛 = 30)
Radmayr et al., Laser Acupuncture Western medicine:
6 months Not reported 13/20 (65%) 15/20 (75%) Response rate, partial response rate
2001 [16] (𝑛 = 20) desmopressin (𝑛 = 20)
5
6
Table 2: Continued.
Duration of Follow-up Experimental Cure rate of Cure rate of
Study Control treatment Outcome measurement
treatment after treatment treatment intervention group control group
Placebo therapy:
Wet nights/wk, voiding frequency,
without active laser
Radvanska et al., Laser acupuncture nocturnal urine production on wet nights
5 weeks Not reported light but with or Not reported not reported
2011 [17] (𝑛 = 16) MVV (maximal voided volume), AVV
without skin contact
(average voided volume)
(𝑛 = 13)
Ear point tapping
Yang et al., 2012 with medicinal Chinese patent
1 month Not reported 21/35 (60%) 12/34 (35.3%) Cure rate, improvement rate
[18] cake-separated medicine (𝑛 = 34)
moxibustion (𝑛 = 35)
Acupuncture-
Chinese medicine
Luo, 2010 [19] 3 months Not reported massage 14/20 (70%) 4/20 (20%) Cure rate, improvement rate
(𝑛 = 20)
(𝑛 = 20)
Tang et al., 2012 Chinese medicine
2 weeks 1 month Massage (𝑛 = 24) 16/24 (66.7%) 11/24 (45.8%) Cure rate, improvement rate
[20] (𝑛 = 24)
Ear point tapping Chinese medicine
Qiu, 2008 [21] 1 month Not reported 17/31 (54.8%) 15/25 (60%) Cure rate, improvement rate
(𝑛 = 31) (𝑛 = 25)
Zhu et al., 2003 Acupoint injection Western medicine:
3 weeks 3 months 19/41 (46.5%) 6/35 (17.1%) Cure rate, improvement rate
[22] (𝑛 = 41) Meclofenoxate (𝑛 = 35)
Medicinal
cake-separated
Western medicine:
Zhang, 2010 [23] 1 month Not reported moxibustion with 22/40 (55%) 6/40 (15%) Cure rate, improvement rate
desmopressin (𝑛 = 40)
embedded needling
(𝑛 = 40)
Chen and Gu, 2003 Acupoint injection Chinese medicine
2 weeks Not reported 36/40 (90%) 14/32 (43.7%) Cure rate, improvement rate
[24] (𝑛 = 40) (𝑛 = 32)
Complete improvement rate, partial
Yuksek et al., 2003 Western medicine:
6 months Not reported Acupressure (𝑛 = 12) 10/12 (83.3%) 7/12 (58.3%) improvement rate, follow-up at 15 days and
[25] oxybutynin (𝑛 = 12)
1, 3, and 6 months
Hong and Zhang, Needle warming Chinese medicine
1 month Not reported 13/15 (86.7%) 5/15 (33.3%) Cure rate, improvement rate
2009 [26] moxibustion (𝑛 = 15) (𝑛 = 15)
reported complete follow-up of all subjects [10, 13, 14, 17]. respectively. The pooled data showed significant difference
All the studies presented selective reporting, characterized between acupuncture therapy and Meclofenoxate (OR = 2.81;
similarity of baseline. In general, the methodological and 95% CI, 1.62–3.96; 𝑃 < 0.0001), with no obvious heterogene-
report qualities of the included studies were poor. ity (Figure 3). The pooled effects of three independent trials
suggested that there was no significant difference between
3.3. Meta-Analysis Results. The 21 included RCTs adopted desmopressin and acupuncture in treating NE (OR = 1.57;
in consistent interventions and different reported outcomes, 95% CI, 0.38–6.57; 𝑃 = 0.54) (Figure 4). Since only one trial
with no unified efficacy standard. To reach a consistent utilized imipramine hydrochloride as medicine control and
understanding of the therapeutic effect of acupuncture ther- only one trial utilized oxybutynin, results from these two
apy for nocturnal enuresis, intervention therapies for control studies are presented as narrative description. There was no
group were further refined. We limited the control group significant difference between imipramine hydrochloride and
methods to western or traditional Chinese medicine alone, acupuncture therapy (OR = 1.71; 95% CI, 0.65–4.51; 𝑃 = 0.27).
as two studies used placebo treatment or sham-acupuncture Compared with oxybutynin, acupuncture could not further
as control group [10, 17] and one of these two studies did not improve the clinical effect (OR = 3.57; 95% CI, 0.53–2; 𝑃 =
report the cure rate as effective outcomes [17]. Furthermore, 0.54).
the definition of cure rate was consistent among the other
included 19 studies; we conducted the meta-analysis to 3.5. Acupuncture Therapy versus Placebo Treatment. Two
compare the overall cure rate determined in these studies. studies used placebo treatment or sham-acupuncture as
Three studies reported mean weekly number of wet nights control group [10, 17]. However, results of these two stud-
[10, 13, 17] and two studies reported maximum voided volume ies were inconsistent. Radvanska et al. [17] compared the
(MVV) [13, 17] as the effective outcomes, considering the lack treatment efficacy of laser acupuncture therapy with sham-
of adequate numbers of studies; these results will be presented acupuncture; they found no significant effect of active laser
in the following part of our review. acupuncture on maximal voided volume (first morning
The results of heterogeneity tests indicated that 𝐼2 > 50% void excluded), maximal morning voided volume, voiding
and 𝑃 < 0.1 for the 19 included studies [6–9, 11–16, 18– frequency, enuresis frequency before and after treatment, or
26] and that the overall heterogeneity existed (𝑃 = 0.002, nocturnal urine production among the patient groups, but it
𝐼2 = 54%). Therefore, a random effects model was used. The resulted in a significant increase in average daytime voided
combined effects of 19 independent trial results showed that volume. There was no effect of skin contact during placebo
acupuncture therapy had further improved the cure rate in laser acupuncture. Radvanska et al. [17] concluded that laser
patients with nocturnal enuresis when compared with control acupuncture had subtle effects on bladder reservoir function;
group accepting medicine therapy (OR = 2.58; 95% CI, 1.84– however, it is an inefficient treatment for monosymptomatic
3.61; 𝑃 < 0.0001) (Figure 2). nocturnal enuresis with reduced maximal voided volume.
Karaman et al. [10] evaluated the effect of laser acupuncture
3.3.1. Acupuncture versus Western Medicine. Our meta- therapy on patients with primary monosymptomatic noc-
analysis of ten studies [6, 8, 9, 11, 13, 16, 22, 23, 25], which turnal enuresis. The mean number of bedwetting episodes
compared acupuncture therapy with traditional Chinese was 1.7 per week 6 months after laser therapy and 3.1 in the
medicine, yielded encouraging effects in favor of acupuncture placebo group. Laser acupuncture therapy was significantly
therapy on nocturnal enuresis (OR = 2.16; 95% CI, 1.31–3.55; more beneficial compared to placebo in terms of complete
𝑃 < 0.01). Heterogeneity between studies existed (𝑃 = 0.03; dryness, partial improvement, and decrease in the mean
𝐼2 = 54%) (Figure 2). number of weekly bedwetting episodes.
3.3.2. Acupuncture versus Traditional Chinese Medicine. The 3.6. Other Outcomes
same findings applied to other ten studies [7, 12, 14, 15, 18–21, 3.6.1. Mean Weekly Number of Wet Nights. Three studies
24, 26], which compared acupuncture therapy with western reported mean weekly number of wet nights [10, 13, 17].
medicine, yielded encouraging effects in favor of acupuncture Moursy et al. [13] reported that the difference of reducing
therapy on nocturnal enuresis (OR = 3.03; 95% CI, 1.88–4.88; the mean weekly number of wet nights in laser acupunc-
𝑃 < 0.01). Heterogeneity between studies existed (𝑃 = 0.01; ture group, desmopressin group, and combination of laser
𝐼2 = 56%) (Figure 2). acupuncture and desmopressin group had no statistical
significance (𝑃 > 0.05). Radvanska et al. [17] found that the
3.4. Subgroup Analyses. A subgroup analysis was conducted difference in the reduction of wet nights was not statistically
to further evaluate the clinical effect of acupuncture therapy significant between laser acupuncture group and placebo
and identify the heterogeneity within western medicine group. Karaman et al. [10] showed that laser acupunc-
group. The western medicine group was divided into four ture therapy was significantly more beneficial compared to
groups according to the medication types. Four studies used placebo in terms of a decrease in the mean number of weekly
Meclofenoxate as control intervention [6, 9, 11, 22], three bedwetting episodes as previously mentioned.
studies used desmopressin as medicine control [13, 16, 23],
and the remaining two studies [8, 25] treated nocturnal 3.6.2. Maximum Voided Volume (MVV). Two studies
children with imipramine hydrochloride and oxybutynin, reported maximum voided volume (MVV) [13, 17] as
Experimental Control Odds ratio Odds ratio

Study or subgroup
Events Total Events Total Weight M-H, random, 95% CI Year M-H, random, 95% CI
1.1.1 Acupuncture therapy versus TCM
Chen and Gu, 2003 36 40 14 32 4.2% 11.57 [3.33, 40.27] 2002
Tian and Zhong, 2008 61 120 47 120 8.1% 1.61 [0.96, 2.68] 2008
Qiu, 2008 17 31 15 25 5.0% 0.81 [0.28, 2.36] 2008
Tong and Zhan, 2009 17 30 10 30 5.1% 2.62 [0.92, 7.46] 2009
Hong and Zhang, 2009 13 15 5 15 2.5% 13.00 [2.07, 81.48] 2009
Luo, 2010 14 20 4 20 3.5% 9.33 [2.18, 39.96] 2010
Hong et al., 2011 19 33 8 33 5.1% 4.24 [1.48, 12.17] 2011
Ling and Chen, 2011 18 30 15 30 5.2% 1.50 [0.54, 4.17] 2011
Hong et al., 2011 20 33 8 33 5.0% 4.81 [1.67, 13.86] 2011
Yang et al., 2012 21 35 12 34 5.4% 2.75 [1.04, 7.30] 2012
Tang et al., 2012 16 24 11 24 4.5% 2.36 [0.73, 7.60] 2012
Subtotal (95% CI) 411 396 53.8% 3.03 [1.88, 4.88]
Total events 252 149

Heterogeneity: 𝜏2 = 0.34; 𝜒2 = 22.66, df = 10 (P = 0.01); I2 = 56%
Test for overall effect: Z = 4.55 (P < 0.00001)
1.1.2 Acupuncture therapy versus western medicine

Radmayr et al., 2001 13 20 15 20 3.8% 0.62 [0.16, 2.43] 2001
Zhu et al., 2003 19 41 6 35 5.0% 4.17 [1.43, 12.20] 2003
Yuksek et al., 2003 10 12 7 12 2.4% 3.57 [0.53, 23.95] 2003
Hui et al., 2006 20 35 14 32 5.5% 1.71 [0.65, 4.51] 2006
Liu, 2007 18 30 9 30 5.0% 3.50 [1.20, 10.20] 2007
Ding et al., 2007 25 42 13 38 5.8% 2.83 [1.14, 7.03] 2007
Zhang, 2010 22 40 6 40 5.0% 6.93 [2.38, 20.16] 2010
Dong et al., 2012 33 60 26 60 6.9% 1.60 [0.78, 3.29] 2012
Moursy et al., 2014 33 62 35 62 6.9% 0.88 [0.43, 1.78] 2014
Subtotal (95% CI) 342 329 46.2% 2.16 [1.31, 3.55]
Test for overall effect: Z = 3.03 (P = 0.002)
Total (95% CI) 753 725 100.0% 2.58 [1.84, 3.61]


0.001 0.1 1 10 1000
Favouring medication Favouring acupuncture
Test for subgroup differences: 𝜒2 = 0.93, df = 1 (P = 0.34), I2 = 0%
Figure 2: Forest plot of comparison: the clinical effective rate.
the effective outcomes. Moursy et al. [13] found that it associated with Egger’s test was 0.002. The resulting graph was
significantly increased only in laser acupuncture group and asymmetrical, suggesting the possibility of publication bias,
combination of laser acupuncture and desmopressin group which was in line with results of Begg’s test and Egger’s test
comparing with pretreatment values and desmopressin (Figure 5). In addition, language bias may exist because most
group, respectively. Thus, bladder capacity significantly of included studies were published in Chinese.
increased only in patients receiving laser acupuncture
treatment. However, Radvanska et al. [17] reported that the 4. Discussion
MVV had no difference between laser acupuncture group
and placebo group. 4.1. Summary of Evidence. The present study analyzed data
from 21 RCTs involving 1590 individuals that featured to
3.7. Publication Bias Analysis. We conducted a funnel plot assess the efficacy of acupuncture therapy to treat NE. Based
analysis of the aforementioned 19 studies [6–9, 11–16, 18–26]. on the findings in our systematic review and meta-analysis,
𝑃 value associated with Begg’s test was 0.009 and 𝑃 value acupuncture therapy can significantly improve the clinical
Experimental Control Odds ratio Odds ratio

Study or subgroup
Events Total Events Total Weight M-H, fixed, 95% CI Year M-H, fixed, 95% CI
Zhu et al., 2003 19 41 6 35 14.3% 4.17 [1.43, 12.20] 2003
Liu, 2007 18 30 9 30 14.8% 3.50 [1.20, 10.20] 2007
Ding et al., 2007 25 42 13 38 22.7% 2.83 [1.14, 7.03] 2007
Dong et al., 2012 33 60 26 60 48.2% 1.60 [0.78, 3.29] 2012
Total (95% CI) 173 163 100.0% 2.53 [1.62, 3.96]

Total events 95 54
Heterogeneity: 𝜒2 = 2.81, df = 3 (P = 0.42); I2 = 0%
0.01 0.1 1 10 100
Favouring Meclofenoxate Favouring acupuncture
Figure 3: Subgroup analysis: acupuncture therapy versus Meclofenoxate.
Experimental Control
Study or subgroup Odds ratio Odds ratio
Events Total Events Total Weight M-H, random, 95% CI Year M-H, random, 95% CI
Radmayr et al., 2001 13 20 15 20 29.7% 0.62 [0.16, 2.43] 2001
Zhang, 2010 22 40 6 40 33.2% 6.93 [2.38, 20.16] 2010
Moursy et al., 2014 33 62 35 62 37.1% 0.88 [0.43, 1.78] 2014
Total (95% CI) 122 122 100.0% 1.57 [0.38, 6.57]

Total events 68 56
0.01 0.1 1 10 100
Test for overall effect: Z = 0.62 (P = 0.54)
Favouring desmopressin Favouring acupuncture
Figure 4: Subgroup analysis: acupuncture therapy versus desmopressin.
0 4.2. Mechanism of Acupuncture Therapy. The pathogenesis of

nocturnal enuresis is multifactorial; several factors such as
0.2 psychosocial, developmental, hormonal, and genetic factors
have been proven to be involved in nocturnal enuresis.
Nocturnal polyuria, nocturnal detrusor overactivity, and high
SE(log[OR])
0.4
arousal thresholds are main pathogenesis of NE. To date,
0.6
increasing evidence suggests that all three mechanisms can
be attributed to brainstem disturbance. The locus coeruleus
(LC) has axonal connections with the hypothalamic cells
0.8
that produce vasopressin and also plays an important role in
arousal from sleep [29, 30]. Pontine micturition center coor-
1 dinates the micturition reflex and overlaps both functionally
0.001 0.1 1 10 1000
and anatomically with LC. A disturbance in this region of
OR
brainstem may cause a range of pathological changes which
Subgroups may result in the pathogenesis of NE.
Acupuncture therapy versus TCM
Acupuncture therapy versus western medicine
Acupuncture points were selected in order to influence
the spinal micturition centers as well as the parasympathetic
Figure 5: Funnel plot of randomized controlled trials. innervation to the urinary tract [31]. With acupuncture
stimulation, levels of enkephalins and endogenous opioids
are increased in both plasma and central nervous system.
efficacy in enuretic children when compared with placebo An increased beta-endorphin level in human cerebrospinal
acupuncture or TCM. In contrast to western medicine, fluid could be detected after acupuncture stimulation [32].
acupuncture therapy was more effective than Meclofenoxate. And beta-endorphin was found to be able to depress bladder
Conclusions regarding the safety of acupuncture therapy contractions [33]. The therapeutic effects of acupuncture
cannot be drawn due to the paucity of evidence provided by therapy can be achieved through the suppression of spinal
the included trials. However, the drawn conclusion should be and supraspinal reflexes which lead to bladder contraction.
interpreted cautiously owing to low methodological qualities And the clinical efficacy of acupuncture was reflected in
of included studies. increase in maximum bladder capacity and suppression of
detrusor muscle activity; these functional changes might terms, cure rate, complete improvement rate, and response
contribute to the improvement of NE. rate, are synonyms; children having no bedwetting episodes
In TCM theory, the generation and discharge of urine on follow-ups were defined to be cured. The majority of
are associated with lung, kidney, spleen, and bladder. The our eligible studies failed to distinguish between NMNE and
pathogenesis of nocturnal enuresis is Qi deficiency of lung, MNE, making it difficult to get a precise conclusion. To our
spleen, and kidney; bladder is not controlled by Qi as well. knowledge, there is still no worldwide unified evaluation
Through different forms of stimulation on meridian points standard to assess the basic state and disease’s progression of
or specific parts of body, imbalance and instability between enuretic children. In addition, the duration of acupuncture
Zangfu organs are corrected to improve symptoms of NE and sessions and follow-ups after treatments vary from studies
maintain the stability of inner state [34]. Based on the classical to studies. Since acupuncture therapy has a long-lasting
prescriptions of acupuncture, series of novel acupuncture beneficial effect on enuretic children, the outcomes were
modalities have been widely applied in clinic. In our sys- supposed to be measured at the end of follow-ups after
tematic review, the specific interventions employed in these treatment.
eligible trials included traditional fine needle acupuncture, The utilization of different acupuncture techniques by
moxibustion, electroacupuncture, auricular point sticking, different investigator can greatly affect curative effect of
acupoint catgut embedding, acupressure, transdermal drugs acupuncture therapy [5]. Based on TCM theory, all acupunc-
delivery systems, and acupoint injection. These techniques ture procedures need to be performed according to syndrome
were considered as one type of therapy, without differen- differentiation. A lack of understanding of TCM was reflected
tiating acupoint selection or acupuncture forms. Therefore, in the treatment models; treatment following the same
the findings in this review might indicate an overall efficacy pattern can reduce the therapeutic effect to some extent.
trend, but definitive conclusions could not be drawn. Acupuncture sessions should be performed based upon strict
diagnosis made by four basic diagnostic methods (inspec-
4.3. Comparison with Other Studies. In 2005, a system- tion, auscultation, olfaction, and palpation) [38]. As various
atic review reported that acupuncture in combination with acupuncture modalities are difficult to master, practitioners
another therapy could further significantly reduce the num- and physicians are required to have a deep understanding
ber of wet nights when compared to acupuncture therapy of the mechanisms underlying NE so that acupuncture
alone, and, regarding the comparison of acupuncture therapy techniques could be applied appropriately. The investigators
with antidiuretic medication, the results showed that the who lack universal knowledge of TCM theory should be
outcome favored medication but was not significantly better encouraged to participate in the standardized training before
than acupuncture therapy [35]. Our meta-analysis managed the application of acupuncture.
to summarize all published RCTs to compare the clinical In contrast to TCM, acupuncture therapy could further
efficacy of acupuncture therapy with pharmacological treat- improve the clinical effect in treating nocturnal children;
ment or placebo treatment. The findings in our meta-analysis no subgroup analysis was made in this group because the
suggested that acupuncture therapy was more effective than acupuncture modalities and Chinese medicine types varied
both western diuretic medication and traditional Chinese from studies to studies. The data extracted from these stud-
medicine, which ran counter to the conclusion in aforemen- ies suggested an overall efficacy trend; the standardization
tioned systematic review. of acupuncture techniques is one problem to be solved
in need. In the subgroup analysis conducted in western
4.4. Limitations. Based on the studies included in our meta- medicine group, acupuncture therapy was more effective
analysis, the methodological qualities were judged to be than Meclofenoxate while no significant difference could be
generally poor, which might limit the value of conclusions detected between acupuncture and imipramine hydrochlo-
about clinical efficacy of acupuncture therapy for treating ride, desmopressin, or oxybutynin. Types and doses of
NE. The vast majority of the included trials failed to describe administered drugs might affect the results of experiment
detailed information about randomization and allocation to a certain extent. Given that the evidence from China
concealment. Lack of blinding procedures in RCTs can also occupies a large proportion, further rigorous experiments
exaggerate the conclusions of these trials. Further assessment within western context are required. Considering all these
of acupuncture therapy needs to be taken by large-scale above factors, the appearance of heterogeneity could be
clinical studies which employ rigorous methodologies. reasonably explained.
The diagnosis and therapeutic evaluation standards
employed by studies, that are performed in China, are mainly 4.5. Suggestion for Future Research. The included studies
in accordance with “Standards for Diagnosis of Syndromes or in our systematic review comprise various methodolog-
Diseases of TCM and Evaluation of the Therapeutic Effect” ical deficiencies, and the findings of the present review
issued by the State Administration of TCM in 1994 [36]. In are somewhat limited due to low methodological qualities.
the studies published in English, the majority of recruited Future randomized controlled trials should employ improved
patients are diagnosed and evaluated according to the “Stan- methodologies and reporting specifications as follows: (1)
dardization and Definition of Lower Urinary Tract Dysfunc- all clinical studies of acupuncture should be registered and
tion in Children” of the International Children’s Continence comply with the revised standards for reporting interventions
Society (ICCS) [37]. To conduct a meta-analysis, the outcome in clinical trials of acupuncture (STRICTA) [39]; (2) the
measure adopted in included RCTs was clinical efficacy. Such sample sizes should be calculated; (3) the generation of
random allocation sequences and allocation concealment [6] H. Dong, Y. Shi, and Z. Chai, “Acupoint injection plus scrapping
should be provided in detail; (4) these studies should be for nocturnal enuresis:an observation of 60 cases,” Journal of
blinded and placebo controlled; (5) the standard of diagnosis New Chinese Medicine, no. 6, pp. 135–136, 2012.
should be unified and widely accepted; (6) the follow-ups [7] J. Hong, L. Liu, J. Hong et al., “Clinical research of moxa-
after treatments are required to be at least 6 months so that moxibustion therapy for infantile enuresis,” in Proceedings of
patients could be revalued; (7) all adverse events associated the 19th National Seminar of the Clinical Branch of China
with acupuncture should be reported and rigorously assessed. Acupuncture and Moxibustion Association, pp. 141–143, Zhangji-
ajie, China, 2011.
[8] J. Hui, Y. Zhao, H. Gao, and T. He, “Heat-producing needling
5. Conclusion for pediatric enuresis: an observation of 35 cases,” Journal of
Pediatrics of Traditional Chinese Medicine, vol. 2, no. 1, pp. 48–
In summary, the results of this study suggest that acupuncture 50, 2006.
therapy demonstrate better clinical efficacy than pharmaco-
[9] J. Liu, “Clinical research of Transdermal drug delivery machine
logical treatment or placebo treatment in treating NE. Due
in treatment of nocturnal enuresis,” Modern Health (Medicine
to the low methodological qualities of included trials, the Innovation Research), vol. 4, no. 5, pp. 45–46, 2007.
findings of current study should be interpreted with caution.
[10] M. I. Karaman, O. Koca, E. V. Küçük, M. Öztürk, M. Güneş, and
Therefore, to further assess the potential beneficial effect of
C. Kaya, “Laser acupuncture therapy for primary monosymp-
acupuncture therapy for NE, additional RCTs with rigorous tomatic nocturnal enuresis,” Journal of Urology, vol. 185, no. 5,
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control, long follow-ups, and strict reporting specification are
[11] L. Ding, D. Du, and L. Han, “Transdermal drug delivery
required.
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Conflict of Interests Health, vol. 19, no. 6, pp. 181–213, 2007.
[12] M. Tong and X. Zhan, “30 African school-age children and ado-
The authors declare that there is no conflict of interests lescents with primary nocturnal enuresis treated by suspended
regarding the publication of this paper. moxibustion,” Journal of External Therapy of Traditional Chinese
Authors’ Contribution [13] E. E. S. Moursy, N. F. Kamel, and A. F. Kaseem, “Combined
laser acupuncture and desmopressin for treating resistant cases
Zheng-tao Lv and Wen Song contributed equally to this of monosymptomatic nocturnal enuresis: a randomized com-
paper. parative study,” Scandinavian Journal of Urology, vol. 48, no. 6,
pp. 559–564, 2014.
[14] P. Tian and Z. Zhong, “Clinical effect of acupuncture &
Acknowledgments moxibustion on kidney Qi deficiency syndrome of children
nocturnal enuresis,” Journal of Modern Clinical Medicine, vol.
This work was supported by Grants from the National Natural 34, no. 4, pp. 276–277, 2008.
Science Foundation of China (no. 81473768; no. 81101927) [15] Q. Ling and X. Chen, “Human’s placenta tissue fluid injection
and Grants from Wuhan Science and Technology Bureau no. at acupoint for treating nocturnal enuresis: a clinical effect
2013060602010280. observation of 30 cases,” Journal of Gansu College of Traditional
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http://dx.doi.org/10.1155/2015/642081
Research Article
Regulation of Neurotrophin-3 and Interleukin-1𝛽 and
Inhibition of Spinal Glial Activation Contribute to the
Analgesic Effect of Electroacupuncture in Chronic Neuropathic
Pain States of Rats
Wenzhan Tu,1 Wansheng Wang,2 Haiyan Xi,3 Rong He,1 Liping Gao,1 and Songhe Jiang1
1
Department of Physical Medicine and Rehabilitation, The Second Affiliated Hospital & Yuying Children’s Hospital of
Wenzhou Medical University, Wenzhou, Zhejiang 325027, China
2
Department of Rehabilitation Medicine, The Affiliated Hospital of Binzhou Medical University, Binzhou, Shandong 256603, China
3
Department of Gynecology, The Affiliated Hospital of Binzhou Medical University, Binzhou, Shandong 256603, China
Correspondence should be addressed to Songhe Jiang; songhe.jiang@gmail.com
Received 10 August 2014; Revised 17 December 2014; Accepted 19 December 2014
Academic Editor: Bing Zhu
Copyright © 2015 Wenzhan Tu et al. This is an open access article distributed under the Creative Commons Attribution License,
Growing evidence indicates that neurotrophin-3, interleukin-1𝛽, and spinal glia are involved in neuropathic pain derived from
dorsal root ganglia to spinal cord. Electroacupuncture is widely accepted to treat chronic pain, but the precise mechanism
underlying the analgesic effect of EA has not been fully demonstrated. In this study, the mechanical withdrawal threshold and
thermal withdrawal latency were recorded. We used immunofluorescence and western blots methods to investigate the effect of
EA on the expression of NT-3 and IL-1𝛽 in DRG and spinal cord of CCI rats; we also examined the expression of spinal GFAP and
OX-42 in spinal cord. In present study, the MWT and TWL of CCI group rats were lower than those in the Sham CCI group rats,
but EA treatment increased the pain thresholds. Furtherly, we found that EA upregulates the expression of NT-3 in DRG and spinal
cord of CCI rats, while EA downregulates the expression of IL-1𝛽. Additionally, immunofluorescence exhibited that CCI-induced
activation of microglia and astrocytes was inhibited significantly by EA treatment. These results demonstrated that the analgesic
effect of EA may be achieved through promoting the neural protection of NT-3 as well as the inhibition of IL-1𝛽 production and
spinal glial activity.
1. Introduction [7], which can prevent the development and maintenance

of thermal hyperalgesia [8]. Additionally, NT-3 delivered
Neuropathic pain, characterized by spontaneous pain, hyper- by exogenous administration has been reported to allevi-
algesia, and allodynia, is often caused by peripheral nerve ate the mechanical hyperalgesia induced by intramuscu-
injury [1, 2]. Such pain is often persistent and poorly treated lar acid injection in transgenic mice [9]. Moreover, NT-3
by existing therapies [3]. Now we know that neuroinflam- can be produced by astrocytes [10] and microglia [11]. All
mation [4], purinergic signaling [5] and some other pain these studies demonstrate the benefit of NT-3 on painful
signaling molecules play key roles in the development of neu- states.
ropathic pain. However, the neural protection and inhibition Interleukin-1𝛽 (IL-1𝛽), a polypeptide proinflammatory
of spinal glia mechanism in treating neuropathic pain need cytokine, plays an important role in modulating neuronal
be given more attention to. excitability in the peripheral nervous systems [12]. It is
Neurotrophin-3 (NT-3), a member of the neurotrophins, released under conditions associated with persistent pain
is a target-derived neurotrophic factor that regulates sen- including inflammatory pain and neuropathic pain [12].
sory neuronal survival and growth [6]. NT-3 is a potent Recent studies suggest that spinal IL-1𝛽 may be produced by
negative modulator of the neuropathic pain state associated glial cells (microglia and astrocytes) in different chronic pain
with chronic constriction injury (CCI) of the sciatic nerve states [13, 14]. Furthermore, there is growing recognition that
spinal glia contributes to the development and maintenance At the time of paw withdrawal, the maximum force was
of central sensitization in chronic pain [15]. recorded. Each rat was tested alternately in 5 min intervals,
Acupuncture has been used for more than 3000 years in and each rat was tested 6 times. Excluding the maximum and
traditional Chinese medicine [16]. Electroacupuncture (EA) minimum forces, the average value was used as the MWT.
is a procedure in which fine needles are inserted into an
individual at discrete points, followed by electrical stimula- 2.4. Thermal Withdrawal Latency (TWL). In order to eval-
tion to relieve pain [17, 18]. In traditional Chinese medicine, uate thermal hyperalgesia, the 37370 Plantar Test Apparatus
Zusanli (ST-36) and Yanglingquan (GB34) are commonly (Ugo-Basile, Milan, Italy) was used to test the TWL. The rats
used in acupuncture to treat neuropathic pain in the waist were placed in a transparent acrylic chamber (17 cm × 11.5 cm
and lower extremities. ST36 is located 5 mm beneath the × 14 cm) and given 20 min of adaptation. The radiant heat was
capitulum fibulae and lateral-posterior to the knee joint and set at 50∘ C and placed to the plantar surface of the hind paw.
GB34 is about 5 mm superior-lateral to ST36. ST36 and GB34 The withdrawal of the paw, indicating the sensation of pain
acupoints distribute near the common peroneal nerve and in the rat, caused the infrared source stop and the reaction
the superficial and deep peroneal nerves. Additionally, some time was recorded. The hind paw was tested alternately at
studies [2, 19] have demonstrated that EA might alleviate 10 min intervals and the cut-off time for heat stimulation was
neuropathic pain behavior of CCI rats and we have reported 40 s. Each rat was tested six times over the course of the
that EA could increase pain thresholds of rats with CCI [20]. experiment. Excluding the maximum and minimum times,
When injury occurred, the nociceptive signals would form the average value was expressed as the TWL.
and come into the DRG neurons and then spinal dorsal horn
following the corresponding nerves. Therefore, the analgesic 2.5. Electroacupuncture (EA) Treatment. In the EA group,
effect of EA at ST36 and GB34 acupoints may be achieved by EA was started on day 7 after the CCI injury [22] and
the regulation of NT-3, IL-1𝛽, and spinal glia in DRG neurons then given daily for the following 7 days; all EA was given
or spinal dorsal horn. between 9:00 and 11:00 a.m every day. The rats were main-
The aim of the present study was to investigate whether tained without anesthesia in an immobilization apparatus
the analgesic effect of EA was associated to following mecha- designed by our laboratory (patent application number:
nism: (1) promote the neural protection of NT-3; (2) the anti- 201110021482.5, State Intellectual Property Office), a system
inflammatory effect by decreasing IL-1𝛽; (3) inhibiting the convenient for acupuncture research and helpful to reduce
activation of spinal glia. stress for experimental rats. At ipsilateral ST-36 and GB-34,
two needles were inserted to a depth of approximately 2-3 mm
2. Materials and Methods and connected to the output terminals of an EA apparatus
(HANS-200E, Jisheng Medical Instruments). The frequency
2.1. Animals. The Institutional Animal Care and Use Com- of stimulation was alternately applied as a square wave at
mittee of Wenzhou Medical University approved all exper- 2/100 Hz, and the intensity of the stimulation was applied for
iments performed in accordance with the guidelines of the 30 min at 2 mA.
International Association for the Study of Pain. Male Sprague
Dawley rats (200–250 g) were used for this study. The rats 2.6. Immunofluorescence. Half of all experimental animals
were randomly divided into 3 groups: Sham CCI group, CCI were taken randomly for immunofluorescence study (𝑛 =
group, and CCI plus EA group. All animals were housed in 6 in each group). On day 14, the rats were deeply anes-
plastic boxes at 22–24∘ C and provided free access to food and thetized using 5% chloral hydrate and perfused with 200 mL
water under a 12/12 h reversed light-dark cycle. normal saline into the aorta, followed by 250 mL of 4%
paraformaldehyde in 0.1 M phosphate buffered saline (PBS,
2.2. Chronic Constriction Injury Model. The CCI model of pH 7.2–7.4). Subsequently, the ipsilateral L4-6 DRGs and
neuropathic pain was chosen based on a previous description whole L4-L5 lumbar spinal cords were removed, postfixed,
[21]. Briefly, after all rats were anesthetized with sodium and replaced with 30% sucrose. Transverse spinal sections
pentobarbital (80 mg/kg, i.p.) and the right sciatic nerve (free-floating, 30 mm) and DRG sections (10 mm) were cut
was exposed at the mid-thigh level, proximal to the sciatic in a cryostat (Leica) and processed for immunofluorescence
trifurcation, four ligature knots (4-0 chromic gut) were [23]. To ensure that immunohistochemical data were com-
loosely tied with 1 mm intervals. In the Sham CCI group, parable between groups, free-floating sections were carefully
the right sciatic nerve was exposed for 2-3 minutes but not processed by immunohistochemistry under the same condi-
ligated. tions (such as the washing times, the incubating time, and
the temperature). Followed by a PBS wash for 5 min, five
2.3. Mechanical Withdrawal Threshold (MWT). In order to times, all sections were sequentially blocked with 10% goat
evaluate mechanical allodynia, the 2392 Electronic von Frey serum albumin, for one hour, in PBS + T (0.3% Triton-
Anesthesiometer (IITC Life Science, USA) was applied to X 100) at room temperature and were incubated overnight
estimate the MWT. All rats were placed individually inside at 4∘ C with different primary antibodies: rabbit polyclonal
a wire mesh-bottom cages (20 cm × 14 cm × 16 cm) and given anti-NT-3 (1 : 200, Santa Cruz, USA), rabbit polyclonal anti-
20 min of adaptation. The probe was positioned below the IL-1𝛽 (1 : 200, Santa Cruz, USA), mouse monoclonal anti-
plantar surface of the paw with von Frey filaments at a range GFAP (astrocyte marker, 1 : 1000, Calbiochem, USA), and
of 0.1–70 g, with increasing force until the rat paw twitches. mouse monoclonal anti-OX-42 (microglia marker, 1 : 1000,
22
30 20
Mechanical withdrawal threshold (g)
Thermal withdraw latency (s)

18
25
16
###
20 14
### 12
15
10
8
10
6
&&& &&&
5 4
0 3 5 7 10 14 0 3 5 7 10 14
Day Day
Sham CCI Sham CCI
CCI CCI
CCI + EA CCI + EA
(a) (b)
Figure 1: Analgesic effects of EA treatment on mechanical withdrawal threshold (MWT) and thermal withdrawal latency (TWL) induced by
chronic constrictive injury. On day 14, the MWT (a) and TWL (b) in each group were recorded and compared with each other. &&& 𝑃 < 0.001,
versus the Sham group; ### 𝑃 < 0.001, versus the CCI group.
ABD Serotec, USA). After the primary antibody incubation, Santa Cruz, USA). The membranes were washed four times
the sections were then incubated for 1 h at room temperature with TBST and incubated (1.5 h, room temperature) with
with secondary antibodies (1 : 400, DyLight 488-labeled goat horseradish peroxidase-conjugated secondary antibody (goat
anti-rabbit IgG or DyLight 594 AffiniPure goat anti-mouse anti-rabbit IgG 1 : 5000, Chemicon, USA) in blocking buffer.
IgG, EarthOx, USA). Finally, sections were washed with PBS After being washed, the labeled proteins were visualized
and the cover-slips were mounted onto slides using antifade using the enhanced chemiluminescence (ECL) kit (Beyotime,
mounting medium (Beyotime, China). The stained sections China). The immune complex was collected on Kodak light
were examined with a fluorescence microscope (Olympus, film and the quantity of band intensity was detected by a
Japan). DNR Micro Chemi Chemiluminescence gel imaging system.
The quantitative analysis was performed on each ani- The band densities were normalized to each glyceraldehyde-
mal from five randomly selected sections per animal. The 3-phosphate dehydrogenase (GAPDH).
immunofluorescence brightness and density of the staining
were tested by Image Pro Plus software: the immunofluo- 2.8. Statistical Analysis. All results were expressed as the
rescence density was used for examining the positive cell of means ± standard deviation (SD). The statistical differences
NT-3 in DRGs; the immunofluorescence bright area was used were analyzed using one-way ANOVA with Tukey or Dun-
for examining the expression of IL-1𝛽, GFAP, and OX-42 in nett’s post hoc tests for multiple comparisons. A 𝑃 value <
spinal cord. 0.05 was considered statistically significant.
2.7. Western Blots. The remainder of experimental animals 3. Results

were used for western blots (𝑛 = 6 in each group). On
day 14 after the CCI operation, the rats were deeply anes- 3.1. Effect of EA on Mechanical and Thermal Hyperalgesia of
thetized and the L4-L5 spinal cord segments were isolated CCI Rats. Before and on days 3, 5, 7, 10, and 14 after the CCI
immediately and flushed with ice-cold PBS. The segments operation, the MWT and TWL were measured. On day 14,
were lysed and microfuged at 12,000 rpm for 5 min at 4∘ C, the MWT and TWL in the CCI group were significant lower
and subsequently the supernatant was collected. Protein than those in the Sham CCI group (𝐹(2,21) = 458.4, 𝑃 < 0.001;
samples (30 𝜇g) were loaded on a 10% Tris–HCl SDS-PAGE 𝐹(2,21) = 144.6, 𝑃 < 0.001). However, the MWT and TWL
gel (Bio-Rad, Hercules, CA) for 30 min at 70 V and 55 min in the EA group were higher than those in CCI group (𝑃 <
at 120 V. After electrophoresis, the proteins were electro- 0.001, 𝑃 < 0.001), implying EA treatment could increase the
transferred to a polyvinylidene fluoride (PVDF) membrane mechanical and thermal threshold in the rats suffering from
for 50 min at 300 mA. The membranes were blocked with neuropathic pain after CCI operation (Figures 1(a) and 1(b)).
Tris-buffered saline (TBS), containing 0.1% Tween-20, 5%
skim milk, and 0.2% BSA for 2 h at room temperature 3.2. Effect of EA on the Immunoreactive Changes of NT-3
and incubated over night at 4∘ C with primary antibodies: in DRGs. The expression of NT-3 in DRG was observed
anti-NT-3 (1 : 300, Santa Cruz, USA) and anti-IL-1𝛽 (1 : 250, through immunofluorescence. The immunofluorescence
Sham CCI CCI CCI + EA
(a)
###
25
Positive neuron in DRG (%)
20
15 &&
10
0
(b)
Figure 2: Effect of EA on CCI-induced increase of the immunoreactive changes of NT-3 in DRG. (a) NT-3 immunopositive neurons in
ipsilateral DRG of each group. (b) Quantification of positive neurons showing that EA treatment promoted the expression of NT-3. && 𝑃 < 0.01,
versus the Sham group; ### 𝑃 < 0.001, versus the CCI group.
density >12 was used for examining positive cells. In the 3.5. Inhibitory Effect of EA on Spinal Glial Activation. By
CCI group, the number of positive neurons was more than means of immunohistochemistry, we used GFAP and OX-
that in the Sham CCI (𝐹(2,15) = 61.1, 𝑃 < 0.001). However, 42 to label astrocyte and microglia in the spinal cord,
after EA treatment, the expression of NT-3 increased further respectively. Expression of GFAP and OX-42 was obviously
(𝑃 < 0.001; versus the CCI group) (Figures 2(a) and 2(b)). upregulated in the spinal dorsal horn on day 14 after CCI
injury (𝐹(2,15) = 44.4, 𝑃 < 0.001 and 𝐹(2,15) = 67.48,
3.3. Effect of EA on the Quantitative Changes of NT-3 Protein 𝑃 < 0.001, versus Sham CCI group). The expression of GFAP
in Spinal Cord. The NT-3 expression at the protein level and OX-42 in CCI + EA group was lower than that in CCI
in spinal cord was analyzed using western blotting. The group (𝑃 = 0.002 and 𝑃 = 0.003). This result indicated that
relative optical density (ROD) value for the NT-3 protein EA could inhibit the activation of astrocyte and microglia
expression in the CCI group was significantly higher than induced by CCI (Figures 6(a) and 6(b)).
that in Sham CCI group (𝐹(2,15) = 96.2, 𝑃 < 0.001), and
EA treatment increased that ROD in the EA group. Similar 4. Discussion
to the result of immunohistochemistry, the level of NT-3
protein significantly increased after EA treatment (𝑃 < 0.001) NT-3 is a target-derived neurotrophic factor that regulates
(Figures 3(a) and 3(b)). sensory neuronal survival and growth [6]. It has been
reported NT-3 could prevent the development and mainte-
3.4. Effect of EA on the Immunoreactive and Quantitative nance of thermal hyperalgesia with CCI of the sciatic nerve
Changes of IL-1𝛽 in Spinal Cord. The expression of IL-1𝛽 in [8]. So, NT-3 may release the neural plasticity caused by CCI
spinal cord was observed through immunofluorescence and and decrease the neurocells sensibility to stimulation. Here
western blotting. CCI injury increased the expression of IL-1𝛽 we have shown that the pain hypersensitivity of CCI rats
in spinal cord (𝐹(2,15) = 50.2, 𝑃 < 0.001; 𝐹(2,15) = 144.6, 𝑃 < was released and NT-3 protein was upregulated in DRG and
0.001), compared with the Sham CCI group (Figures 4 and 5). spinal cord after EA treatment. These findings suggest that
However, in the CCI + EA group, the immunoreactivity and NT-3 could promote the analgesia effect of EA in neuropathic
quantitativeness of protein of IL-1𝛽 were lower than those in pain.
the CCI group (𝑃 < 0.001, 𝑃 < 0.001), implying EA could A growing body of evidence implicates that spinal glia
inhibit the expression of IL-1𝛽 in spinal cord of CCI rats. was involved in the modulation of chronic pain [24] and
140
&&&
120
100
Ratio to GAPDH (%)

80 ###
60
NT-3
40
20
GAPDH
0
Sham CCI CCI CCI + EA Sham CCI CCI CCI + EA
(a) (b)
Figure 3: Effect of EA on CCI-induced increase of the quantitative changes of NT-3 protein in spinal cord. (a) Changes in the relative content
of NT-3 protein in spinal cord of every group. (b) Quantification of bands showing that EA treatment promoted the expression of NT-3.
###
𝑃 < 0.001, versus the Sham group; &&& 𝑃 < 0.001, versus the CCI group.
(a)
10 ###
9
8
7
6
IL-1𝛽
5 &&&
4
3
2
1
0
(b)
Figure 4: Effect of EA on CCI-induced increase of the immunoreactive of IL-1𝛽 in spinal cord. (a) Results of IL-1𝛽 immunofluorescence in
ipsilateral spinal dorsal horn. (b) Changes of the positive area showing that EA treatment suppressed the expression of IL-1𝛽. ### 𝑃 < 0.01,
versus the Sham group; &&& 𝑃 = 0.001, versus the CCI group.
EA analgesia [15, 25]. Painful syndromes are associated with reaction, whereas EA inhibits the reaction. Furthermore, it
different glial activation states: glial reaction (i.e., upregu- has been demonstrated that knockdown of NT-3 markedly
lation of glial markers such as glial fibrillary acidic protein increased the expression of GFAP, OX-42 in the spinal dorsal
(GFAP) and OX-42 and/or morphological changes, including horn during inflammatory pain [15]. This finding suggests
hypertrophy and proliferation) [26]. In parallel with these that the antihyperalgesic role of NT-3 in neuropathic pain
reports, the present study showed that CCI promotes the glial may be mediated through the inhibition of glial activity.
60
###
50
Ratio to GAPDH (%)

40 &&&
30
20
IL-1𝛽
10
GAPDH
0
Sham CCI CCI CCI + EA Sham CCI CCI CCI + EA
(a) (b)
Figure 5: Effect of EA on the increase of IL-1𝛽 protein induced by CCI in spinal cord. (a) Results of western blot from spinal cord in each
group. (b) Changes in the relative content of IL-1𝛽 protein from picture (a). ### 𝑃 < 0.01, compared with the Sham group; &&& 𝑃 < 0.001,
compared with the CCI group.
Spontaneous and evoked pain after nerve injury are findings, we thought that IL-1𝛽 might increase the expression
thought to derive from hyperexcitability of primary and/or of NT-3 at the start of neuropathic pain, while NT-3 might
secondary afferent neurons generated by neurotrophins and depress the expression of IL-1𝛽 at the following stage. Indeed,
proinflammatory cytokines released from activated inflam- EA treatment was given on day 7 after CCI injury in this
matory cells including microglia [27, 28] that become acti- study. So, EA may have an anti-inflammatory effect through
vated in the dorsal horn, astrocytes [29], and macrophages upregulating the expression of NT-3. This conclusion is in
that invade the lesion site [30]. IL-1𝛽 is a cytokine released accordance with the study that demonstrated that NT-3 might
from spinal glial cells in response to pathophysiological serve as an anti-inflammatory factor to suppress neuropathic
changes that occur during different disease states, such as pain [39].
neuropathic pain and inflammatory [12, 31]. Initial reports The relationship between NT-3, IL-1𝛽, and spinal glial
suggested that IL-1𝛽 is an extremely potent hyperalgesic cell is very important to explain the analgesic effect of EA in
agent when injected systemically, intraperitoneally, or intra- neuropathic pain states of rats now. Recently, it was reported
plantarly in rats [32]. In addition, the hypothalamic and that antisense oligodeoxynucleotides specifically against NT-
ventral midbrains mRNA levels of IL-1𝛽 raised by inflam- 3 intrathecally administered could suppress expression of
mation could be reversed to normal levels by acupuncture spinal GFAP, OX-42, and proinflammatory cytokines stim-
stimulation [33]. Now that glia plays an important role ulated with arthritis [15]. In addition, the inhibition of
in nociceptive transmission in neuropathic pain [34] and proinflammatory mediators by NT-3 pretreatment in primary
glial activity would be inhibited by NT-3; the synthesis and microglia with LPS stimulation was corroborated [40]. Based
secretion of proinflammatory cytokines from glial cells would on these published reports and the result of our study, we
be decreased too. Corresponding with this conclusion, we thought NT-3 may be involved in the analgesic effect of
also found the upregulation of IL-1𝛽 in spinal cord in CCI EA on neuropathic pain states of rats mediated through the
rats was suppressed after EA treatment. inhibition IL-1𝛽 production and spinal glial activity.
In recent years, we have changed it as follow: more and The perception of pain requires the activation of multiple
more attention given to the possible roles of neurotrophins neurons across the pain system and the interactions between
and cytokines in the therapeutic effects of acupuncture. The the thalamus, cortex, and limbic system [41]. When injury
cross talk of neurotrophins and cytokines from peripheral occurs, the peripheric receptor is activated and nociceptive
nervous system (PNS) to central nervous system (CNS) is signals are carried from the periphery to the dorsal horn of
involved in the pathophysiology of many human diseases the spinal cord mostly by two populations of small diameter
and may contribute to the effects of acupuncture [35]. primary afferents, the peptidergic and the nonpeptidergic
Therefore, the relationship between NT-3 and IL-1𝛽 may [42]. The peptidergic population expresses neuropeptides,
be an important contributor to chronic pain mechanism. It such as substance P and calcitonin gene-related peptide,
has been reported that IL-1𝛽 could act on sensory neurons while the nonpeptidergic fibers are devoid of neuropeptides,
to increase their susceptibility for injury [36, 37], while express the purinergic receptor P2X3 , and bind the isolectin
NT-3 might release the neural plasticity caused by nerve B4 (IB4) [42]. Considerable studies have demonstrated that
injury and decrease the neurocells sensibility to stimulation. chronic constriction injury of the sciatic nerve induces
Interestingly, it has been demonstrated that IL-1𝛽 could persistent pain behaviors in rats [43, 44] and purinergic
upregulate the expression of NT-3 [38]. Based on these signaling has been proved to be implicated in neuropathic
GFAP
OX-42
(a)
20 ###
GFAP and OX-42 positive area (%)
18
###
16
14
&&
12
10 &&
8
6
4
2
0
GFAP
OX-42
(b)
Figure 6: Effect of EA on CCI-induced increase of the immunoreactivity of GFAP and OX-42 in the spinal dorsal horn. (a) Results of
GFAP and OX-42 immunofluorescence in ipsilateral spinal dorsal horn. (b) Changes of the positive area from picture (a) showing that EA
treatment suppressed the expression of GFAP and OX-42 in spinal cord. ### 𝑃 < 0.001 and ### 𝑃 < 0.001, versus the Sham group; && 𝑃 = 0.002
and && 𝑃 = 0.003, versus the CCI group.
pain [43, 45, 46]. Additionally, our previous study [20] has is a novel finding with respect to modulation of neuropathic
proved that EA might increase the pain thresholds through pain [48]. In addition, the involvement of C-type afferents in
downregulating the expression of P2X3 receptor. It is a truth EA analgesia has been proved [17]. Therefore, we speculated
that some purinergic receptors (e.g., P2X4 and P2X7 ) are that NT-3 may depress purinergic signals in the analgesic
coexpressed with spinal glial. So, following the inhibition of effect of EA, which needs further study.
glial activity, purinergic signaling would be inhibited too. A Based on previous [20] and the present study, we thought
previous study has also shown that peripheral P2X receptors that the neuroprotective effect of NT-3 plays a key role in
are involved in mediating the peripheral excitation of C- and the analgesic effect of EA. The incremental NT-3 inhibited
A𝛿-fiber [47]. Importantly, the ability of NT-3 to prevent and the activation states of spinal glia and downregulated the
reverse thermal hyperalgesia, believed signaled by C-fibers, expression of IL-1𝛽. All these changes promote the stability of
neurocells and decrease the generation of pain signals which [8] T. D. Wilson-Gerwing, C. L. Stucky, G. W. McComb, and V. M.
is transmitted through C-fiber by purinergic signaling. K. Verge, “Neurotrophin-3 significantly reduces sodium chan-
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Conflict of Interests hippocampal astrocytes,” European Journal of Neuroscience, vol.
4, no. 6, pp. 459–471, 1992.
The authors declare that there is no conflict of interests
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http://dx.doi.org/10.1155/2015/210120
Research Article
Brain Network Response to Acupuncture Stimuli in
Experimental Acute Low Back Pain: An fMRI Study
Yu Shi,1 Ziping Liu,1 Shanshan Zhang,1 Qiang Li,2 Shigui Guo,1

Jiangming Yang,3 and Wen Wu1
1
Department of Rehabilitation, Zhujiang Hospital, Southern Medical University, Guangzhou 510282, China
2
Department of General Surgery, Zhujiang Hospital, Southern Medical University, Guangzhou 510282, China
3
Department of Radiology, Zhujiang Hospital, Southern Medical University, Guangzhou 510282, China
Correspondence should be addressed to Wen Wu; wuwen66@163.com
Received 26 November 2014; Revised 2 May 2015; Accepted 5 May 2015
Copyright © 2015 Yu Shi et al. This is an open access article distributed under the Creative Commons Attribution License, which
Most neuroimaging studies have demonstrated that acupuncture can significantly modulate brain activation patterns in healthy
subjects, while only a few studies have examined clinical pain. In the current study, we combined an experimental acute low
back pain (ALBP) model and functional magnetic resonance imaging (fMRI) to explore the neural mechanisms of acupuncture
analgesia. All ALBP subjects first underwent two resting state fMRI scans at baseline and during a painful episode and then
underwent two additional fMRI scans, once during acupuncture stimulation (ACUP) and once during tactile stimulation (SHAM)
pseudorandomly, at the BL40 acupoint. Our results showed that, compared with the baseline, the pain state had higher regional
homogeneity (ReHo) values in the pain matrix, limbic system, and default mode network (DMN) and lower ReHo values in frontal
gyrus and temporal gyrus; compared with the OFF status, ACUP yielded broad deactivation in subjects, including nearly all of
the limbic system, pain status, and DMN, and also evoked numerous activations in the attentional and somatosensory systems;
compared with SHAM, we found that ACUP induced more deactivations and fewer activations in the subjects. Multiple brain
networks play crucial roles in acupuncture analgesia, suggesting that ACUP exceeds a somatosensory-guided mind-body therapy
for ALBP.
1. Introduction gray (PAG), prefrontal cortex] and somatosensory system

(thalamus, primary somatosensory cortex (S1), secondary
Low back pain (LBP) is one of the most common clinical syn- somatosensory cortex (S2), posterior parietal cortices, insula,
dromes and affects 80–85% of people at some point in their supplementary motor area, striatum, and cerebellum) areas
life. This disorder typically causes serious socioeconomic [7, 8], as well as the pain matrix (S1, S2, insular, frontal lobe
problems, including health and economic issues, and even and parietal lobe). The pain matrix showed a strong relation-
medications abuse [1, 2]. Most LBP does not have a definitive ship with pain, which plays an important role in the con-
cause, and it has been shown that traditional Chinese medical duction and communication of pain [9]. Moreover, research
acupuncture has beneficial effects on this intractable pain on acupuncture analgesia has recently become increas-
[3, 4]. ingly popular. Some researchers have found that acupunc-
Previous brain imaging studies have found that external ture yields activations in the attentional- and emotional-
stimuli, including acute experimental pain, could evoke related regions (DMN, dorsomedial, and dorsolateral pre-
deactivations in the default mode network (DMN), a network frontal cortex (dmPFC and dlPFC)), and deactivations in
believed to be involved in the areas of memory and social the somatosensory system (left anterior insula, bilateral S1,
affective and self-referential cognition [5, 6]. Pain stimu- and S2) compared with cutaneous stimuli. Therefore, some
lus could also induce extensive activations in the limbic researchers considered that acupuncture could function as a
system [anterior cingulated cortex (ACC), periaqueductal somatosensory-guided mind-body therapy [10], while others
held the belief that the effect of acupuncture may not be lim-
ited to DMN or the somatosensory system. These researchers
found that acupuncture evoked more deactivations of the
limbic-paralimbic-neocortical network, which was thought BL40 ALBP model location
to be centered on the limbic system, but fewer activations in
the somatosensory and attentional systems compared with
cutaneous stimuli [11]. Taken together, these fMRI studies, Figure 1: The location of the BL40 and ALBP model.
with or without experimental heat pain on limbs, have greatly
contributed to our understanding of the analgesic mechanism
of acupuncture; however, only a few have examined clinical
pain, with the least focus on acute low back pain (ALBP).
Hence, it is interesting to explore how acupuncture modulates
the brain networks in ALBP subjects using fMRI.
There are two major barriers preventing fMRI studies
on clinical ALBP. Firstly, it is hard to distinguish ALBP
qualitatively and quantitatively from the multiple potential
etiologies and their various degrees. Moreover, it is difficult
to conduct experiments because ALBP is characterized by
sudden onset and aggravation. In addition, using experimen- Figure 2: The posture of the subjects when inserting the needle at
tal heat pain to simulate ALBP is problematic, because it is the point.
difficult to expose the volunteers’ back when they are lying
down in the MRI scanner.
In this block design fMRI study, we introduced a simple
and quantitative ALBP model induced by hypertonic saline transverse crease of the popliteal fossa (Figure 1); therefore,
injection in healthy volunteers to investigate the neural each subject’s keen was leaned on mattress to keep lower
mechanism of acupuncture analgesia. For each experimental limbs in a valgus position for therapeutic stimuli (Figure 2).
ALBP subject, we delivered comparable therapeutic stim-
uli, including ACUP and SHAM at the BL40 acupoint 2.2. Experimental Procedures. Anatomical scans of the brain
(Weizhong) on the right lower limb. Moreover, before and and functional images of sensory control stimulation were
after the application of therapeutic stimuli, we collected data collected prior to stimulation imaging. Initially, the subjects
on subjective pretreatment and posttreatment LBP and their were subjected to a baseline (the normal time) resting state
corresponding acupuncture sensations. (rs) MRI scan for 6 min. A preliminary ALBP model was
Therefore, experimental ALBP could not only act as a induced in the right lower back muscle of each subject using
clinical LBP but also as evaluation criteria for the therapeutic a method modified from previous studies [13, 14]. In the
stimuli. experimental ALBP model, we could control the levels of
pain, which gave the subjects a similar level of pain in the
2. Materials and Methods experiment, resulting in a smaller margin of error. The variety
of clinical LBP cannot meet this requirement.
2.1. Subjects and Acupoint. Twenty-eight healthy, right- After the baseline scan, we located an injection point
handed subjects (11 women; age range: 22–30 years) partici- 2 cm lateral to the spinous process of the fourth lumbar
pated in this study. All participants had some knowledge of vertebra for the ALBP model. Thereafter, we filled an in-
acupuncture due to previous cultural exposure; had never dwelling needle (24 gauge) with sterile hypertonic saline
received acupuncture treatment; had a body mass index (10 mL, 5%) and attached it via a long connecting tube
within the standard range (±10%); had no psychiatric or to a computer-controlled power injector (Spectris Solaris
medical illnesses (i.e., multiple sclerosis and epilepsy); and EP; Medrad, Inc., Warrendale, PA, USA), before vertically
had no painful episodes (including dysmenorrhea) or did not inserting it into the above-described location at a depth of
take any drugs (i.e., antipyretics and sleeping pills) within the 1.5 cm (Figures 1 and 3). After 1 min, the hypertonic saline
last month. The study was conducted with the understanding was injected intramuscularly from the above-mentioned
and written consent of each subject. All experiments and pro- computer-controlled power injector into the ALBP subject.
tocols were approved by the Ethics Committee of Zhujiang This injection included a bolus injection (0.1 mL within
Hospital Affiliated to Southern Medical University, China. 5 s) and subsequent continuous injection (0.15 mL/min) to
In traditional Chinese medicine, BL40 (Weizhong acu- produce persistent ALBP. During the first 6 min of ALBP
point) is considered as one of the four most important acustimulation, we performed an rs-fMRI scan to evaluate the
points and proven to have unique efficacy in the treatment of pain status. After the pain rsfMRI scan, we obtained two
ALBP. For instance, A Complete Collection of Acupuncture functional scans for each ALBP subject: one scan during
and Moxibustion, written by Xufeng who is an acupuncture- acupuncture stimulation (ACUP) and one scan during tactile
moxibustion expert in the Ming Dynasty, states that lumbar- stimulation (SHAM) pseudorandomly, with ALBP occurring
back problems could be treated by puncturing this acupoint continually throughout the scanning process. The ACUP or
[12]. BL40 is anatomically located at the midpoint of the SHAM run comprised a block design with six 30 s blocks of
Experimental procedure for ALBP subject = moderate, 7–9 = strong, and 10 = unbearable) [16]. More-
over, each ALBP subject was asked to rate the intensity of LBP
ALBP model before and after each MRI scan using a 10-point visual analog
Two functional scale (0 = none, 1–3 = mild, 4–6 = moderate, 7–9 = strong,
Anatomic scan Baseline scan Pain status scan scans for ACUP and 10 = unbearable). Correspondingly, the scores of the de qi
and SHAM
sensations were compared between ACUP and SHAM in the
ALBP group [18] and pre- and posttreatment LBP between
6 min 6 min Each for 6 min ACUP and SHAM in the ALBP group, using the Wilcoxon
signed-rank test; 𝑃 values < 0.05 were considered to be statis-
Figure 3: The experimental paradigm for the ALBP subjects
included five steps.
tically significant (SPSS 13.0, IBM Corporation, NY, USA).
2.5. Imaging Data Collection and Analysis. Structural and

rest time (OFF block) interspersed between six 30 s blocks of functional scans were acquired with a 3.0 T Philips Achieva
stimulation (ON block); ACUP (or SHAM) was administered MRI System (Royal Philips Electronics, Eindhoven, Nether-
at BL40 by the same experienced acupuncturist during the lands) with an 8-channel head array coil equipped for
six ON blocks of each functional scan. Each functional scan echo planar imaging. The images were axial and parallel to
lasted for 6 min, and the time interval between the two the anterior commissure-posterior commissure line, which
functional scans (ACUP and SHAM) was set at 20 min to covered the whole brain. Structural images were collected
maximize washout of the sustained effects induced by the prior to functional imaging using a T1-weighted fast spin echo
former therapeutic stimulation (Figure 4). All MRI scans sequence (repetition time/echo time = 500/14 ms, flip angle
were performed with each subject laying still in a Philips = 90∘ , 0.859 mm × 0.859 mm in-plane resolution, slice thick-
3.0 T Achieva scanner (Royal Philips Electronics, Eindhoven, ness = 1 mm). Blood oxygenation level-dependent functional
Netherlands) with their eyes and ears covered. imaging was acquired using a T2∗ -weighted, single-shot,
Notably, we had examined the effects of different injec- gradient-recalled echo planar imaging sequence (repetition
tion speeds (0.1 mL/min, 0.15 mL/min, 0.2 mL/min) of the time/echo time = 2000/40 ms, flip angle = 90∘ , 3.4 mm ×
hypertonic saline after a bolus injection (0.1 mL within 5 s), 3.4 mm in-plane resolution, 180 time points for a total of 360
in the preliminary experiment. We found that 0.15 mL/min seconds). In addition, fMRI image collection was preceded by
was most suitable one for our experiment, because it induced 5 dummy scans to minimize gradient distortion.
a persistent moderate-high pain in ALBP subject.
2.5.1. Preprocessing of Experimental MRI Data. Data analysis
2.3. Acupuncture Modulation. ACUP was administered by
was performed with SPM8 software (http://www.fil.ion.ucl.ac
inserting a nonmagnetic (pure silver), 0.4 mm-diameter,
.uk/spm/). Preprocessing includes motion correction, slice-
60 mm-long acupuncture needle (Beijing Zhongyan Taihe
timing correction, normalization to the Montreal Neurologi-
Medicine Co., Ltd, Beijing, China) vertically into BL40
cal Institute standard brain (MNI152), and spatial smoothing
at a depth of approximately 2 cm (Figure 1). To obtain a
with a Gaussian kernel of full width at half maximum of
subjective acupuncture sensation, namely, de qi sensation
8 mm. For motion correction, the subject’s data was excluded
[15], the needle was manually twirled (±180∘ ) at 1 Hz with
if translation or rotation of the subject’s head movements was
“even reinforcing and reducing” needle manipulation in
more than 1.5 mm or 1.5∘ .
traditional Chinese medicine, while SHAM was delivered
with a von Frey monofilament. The acupuncturist poked this (1) Rs-fMRI Data Analysis. The preprocessing data were
monofilament through a needle-guide tube and tapped it then processed to produce regional homogeneity (ReHo)
gently over the skin of the BL40 with the same amplitude and map image files. The ReHo analysis was performed accord-
rate as that used during ACUP [16]. ing to previous reports [18] and calculated using Kendall’s
Considering that SHAM may cause subjective bias coefficient to measure ReHo or the similarity of a ranked
towards the stimulation, all subjects were asked to keep time series from a given voxel with that of its nearest 26
their eyes and ears closed in order to prevent them from neighboring voxels in a voxelwise manner. Kendall’s coeffi-
discriminating the therapeutic stimulation. Moreover, all cient value was calculated for this voxel, and an individual
subjects were purposely misguided that they would receive Kendall’s coefficient map was obtained for each subject. Each
two different forms of acupuncture and needed to con- ReHo map was divided by its own mean ReHo within the
centrate on the degree of acupuncture sensations of BL40. mask for standardization purposes [18]. The ReHo value
Therefore, SHAM aimed to control for not only the superficial differences between the pain status and baseline were cal-
and cutaneous somatosensory effects around BL40 but also culated using two-tailed, paired 𝑡-tests (𝑃 < 0.05) and cor-
the cognitive processing induced by the subject’s expectation rected for multiple comparisons false discovery rate (FDR).
of “ACUP” [17]. The results were displayed using BrainNet viewer software
(http://www.nitrc.org/projects/bnv/).
2.4. Psychophysical Data Collection and Analysis. After each
MRI scan, each subject was asked to quantify the de qi sensa- (2) Task fMRI Data Analysis. In the first-level analysis, the
tions at BL40 using a 10-point scale (0 = none, 1–3 = mild, 4–6 preprocessing task functional data were modeled using a
ON ON ON ON ON ON ON ON ON ON ON ON
OFF OFF OFF OFF OFF OFF OFF OFF OFF OFF OFF OFF
30 s 30 s 20 min
Figure 4: Each functional scan lasted for 6 min, including six OFF-ON blocks; the time interval between the two functional scans was 20 min.
During the six ON blocks of each functional scan, ACUP or SHAM was applied at BL40.
Low back pain in ALBP subject Primary de qi sensations of ALBP subject

10 10
8 8
6 6
4 4
2 2
0 0
Pretreatment Posttreatment Soreness Numbness Heaviness Fullness
ACUP ACUP
SHAM SHAM
Figure 5: Results of psychophysical analysis in ACUP and SHAM. For ALBP subjects, there were significant differences between the ACUP
and SHAM in the mean value of posttreatment pain (𝑃 = 0.043), soreness (𝑃 = 0.014), and fullness (𝑃 = 0.001).
general linear model. Explanatory variables, including the (S.D. = 0.98) and 5.60 (S.D. = 1.24) and those of posttreatment
stimulation task (ACUP or SHAM, ON status) and the OFF LBP were 3.47 (S.D. = 0.75) and 4.51 (S.D. = 1.06) for ACUP
status, were modeled using a boxcar function that convolved and SHAM, respectively. There were significant differences in
with the canonical hemodynamic response function in SPM8. the score for the soreness and fullness between ACUP and
Subsequently, parameter estimates were assessed using least- SHAM for ALBP subjects (Figure 5).
square regression analyses. Next, statistical parametric maps
of the stimulation task (ACUP or SHAM) minus the OFF 3.2. fMRI Results. Compared with baseline (the normal
status contrast were collected at each voxel for each subject. In time), the pain status showed higher ReHo values in the right
the second-level analysis, a one-sample 𝑡-test was applied to medial prefrontal cortex (mPFC), right middle frontal gyrus,
ACUP (or SHAM) minus the OFF status to assess the main right insula, right precuneus (PCN), right parahippocampus
effect of the stimulation, and a paired 𝑡-test was applied to (PHP), and right posterior lobe-cerebellar tonsil. However,
ACUP minus SHAM to assess differences between the ACUP the pain status showed lower ReHo values in the right
and SHAM conditions in the ALBP subjects. The threshold superior temporal gyrus, left middle temporal gyrus, left S1,
was set (𝑃 < 0.05) and corrected for multiple comparisons left ACC, left PHP, and right inferior parietal lobule (𝑃 < 0.05,
(FDR: <0.05). The resulting images were displayed using rest FDR < 0.05, Table 1, Figure 6).
software (http://restfmri.net/forum/rest). Compared with the OFF status, ACUP significantly
affected the activations and deactivations; deactivations were
3. Result found in the somatosensory system (left primary motor
cortex (M1), S2, and frontal eye field), limbic system (left
3.1. Psychophysical Responses. The intensity of the lower back insula and mammillary body, right hippocampus (HP), bilat-
pain and de qi sensations are expressed below as mean ± stan- eral dmPFC, pregenual ACC (pACC), PAG, and PHP), pain
dard deviation. Soreness, numbness, fullness, and heaviness matrix (left S1, left insular, temporal lobe, and frontal lobe),
were the primary de qi sensations in the current study. In the DMN (right angular gyrus, supramarginal gyrus, lateral tem-
ALBP group, the mean values of pretreatment LBP were 5.40 poral cortex, HP, bilateral dmPFC, and PHP), and bilateral
Table 1: Resting state regional homogeneity alterations corresponding to pain status (pain status compared with baseline) paired 𝑡-test analysis
𝑃 < 0.05, FDR < 0.05.
Peak MNI coordinate
BA Cluster sizes Peak 𝑍-score
𝑋 𝑌 𝑍
R mPFC 8 32 3.02 20 27 60
R middle frontal gyrus 9 211 5.87 3 50 22
L middle temporal gyrus 21 54 −3.49 −70 −54 5
R superior temporal gyrus 38 32 −3.59 55 12 −30
L S1 2 59 −2.88 −60 −20 42
R inferior parietal lobule 40 20 −5.94 66 −36 20
L PHP — 31 −2.62 −10 −3 −21
R PHP 35 29 4.51 30 −7 −21
L anterior cingulate cortex 32 42 −2.74 −6 25 39
R precuneus 7 54 2.70 19 −66 33
R insula 13 60 2.47 39 0 22
R cerebellar tonsil — 53 2.42 11 −60 −48
FDR: false discovery rate; MNI: Montreal Neurological Institute; mPMC: medial prefrontal cortex; PHP: parahippocampus; S1: primary somatosensory cortex.
6.0
L R
−6.5
6.0
−6.5
Figure 6: The brain network change in the pain status (pain status and baseline paired 𝑡-test).
thalamus. The activations, including the right M1, S1, and angular gyrus, supramarginal gyrus, PCN, PHP, HP, and
bilateral supplementary motor areas, right insula, and pMCC, temporal pole), bilateral thalamus, cerebellum anterior lobe,
were limited (𝑃 < 0.05, FDR < 0.05, Table 2, Figures 7 and 8). and lateral occipital gyrus (𝑃 < 0.05, FDR < 0.05, Table 3,
Compared with the OFF status, SHAM only produced Figures 7 and 8).
limited deactivations, such as those in the left insula, left Compared with SHAM, ACUP only produced limited
frontal operculum, and left M1, while widespread activations activations, including those in the right insula and right
included the somatosensory system (right frontal eye field M1. In contrast, widespread deactivations were observed,
and bilateral supplementary motor area), attentional system including those in the somatosensory system (left supple-
(bilateral dlPFC), limbic system (right frontopolar area, mentary motor area, bilateral frontal eye field), attentional
bilateral orbitofrontal cortex, PCN, PHP, HP, temporal pole, system (right dlPFC), limbic system (left PAG, bilateral
amygdala, mammillary body, and PAG), DMN (bilateral pACC, dmPFC, PHP, HP, and mammillary body), DMN
Table 2: fMRI signal changes evoked by ACUP (ACUP (ON status compared with OFF status)) one-sample 𝑡-test analysis 𝑃 < 0.05, FDR <
0.05.
Peak MNI coordinate
𝑋 𝑌 𝑍
Left insula 13 52 −3.51 −42 −15 15
Left M1 6 48 −2.56 −36 10 10
Left S2 43 36 −3.01 −37 −12 2
Left frontal eye field 8 66 −3.26 −15 35 53
Left dlPFC 46 97 3.81 −39 36 18
Right M1 6 348 6.16 51 6 12
Left PAG 44 −3.26 −3 −30 −3
Left PHP 35 −3.02 0 −24 0
Left thalamus 26 −2.56 4 −30 −5
Right dmPFC 8 43 −3.98 15 33 45
Right supramarginal gyrus 40 55 4.87 63 −27 33
Right S1 2 40 3.97 55 −20 30
Right supramarginal gyrus 40 48 −3.28 54 −60 39
Right angular gyrus 39 40 −3.00 50 −60 30
Right lateral temporal cortex 21 34 −2.56 49 −55 26
Right pMCC 31 101 3.98 18 −24 39
Bilateral SMA 79 2.54 12 0 60
Right PHP 35 46 −4.00 24 −27 −18
Right HP 40 −3.89 25 −20 −20
Bilateral pACC 32 34 −3.42 0 33 21
Left dmPFC 24 30 −3.23 15 20 20
FDR: false discovery rate; MNI: Montreal Neurological Institute; M1: primary motor cortex; S2: secondary somatosensory cortex; dlPFC: dorsolateral prefrontal
cortex, periaqueductal grey (PAG); PHP: parahippocampus; dmPFC: dorsomedial prefrontal cortex; S1: primary somatosensory cortex; pMCC: posterior mid-
cingulate cortex; SMA: supplementary motor area; HP: hippocampus; pACC: pregenual anterior cingulate cortex.
(right supramarginal gyrus, angular gyrus, bilateral dmPFC, showed that multiple brain networks play important roles in
PHP, and HP), bilateral thalamus, cerebellar anterior lobe, modulating ALBP.
and lateral occipital gyrus (𝑃 < 0.05, FDR < 0.05, Table 4,
Figures 7 and 8). 4.1. The Network Change in the Pain Status. Similar to other
pain stimulation research, the results indicated higher ReHo
4. Discussion values in some areas of the brain network. The right mPFC,
right middle frontal gyrus, right insula, and right PCN are
To the best of our knowledge, this is the first fMRI study to included in the pain matrix, which has a strong relationship
investigate how acupuncture modulates the brain networks with pain. Different parts of the matrix play different roles in
in experimental ALBP subjects. Behaviorally, we delivered the generation and transmission of pain; for example, S1 and
similar pain to every subject in accordance with the ALBP S2 are associated with algesthesia, while the insular cortex
model, whereas, compared with SHAM, ACUP showed and anterior cingulate are associated with the emotional
stronger acupuncture sensations and weaker pain sensations, component of pain [9]. The higher ReHo values in the
suggesting that acupuncture alleviated ALBP. As previously pain matrix represented the pain state via the ALBP model.
found, our fMRI analysis showed that ACUP induced more The mPFC is associated with the processing of emotional
deactivations but less activations compared with pain status information and mediates the functional interactions among
and SHAM. Furthermore, these deactivations in the ALBP the brain regions that participate in pain processing [19, 20],
subjects were mostly in the regions of the limbic system and whereas PCN is likely involved in the shifting of attention
DMN, including the antinociceptive and affective (pACC, between different spatial locations [21]. Therefore, changes
PAG, aMCC, mammillary body, and dmPFC) and memory in ReHo may reflect pain accompanied by the processing of
(DMN and mammillary body) related brain regions [16]. In emotionally intense information.
contrast, the activations in the ALBP subjects were found ACC participates in pain perception and integration of
in the attentional (dmPFC, dlPFC, pMCC, and right insula) the sensory, attentional, and cognitive components of pain
and somatosensory system (right S1, M1, and insula) related [22, 23]. The decrease of ReHo in ACC suggests a reduction
regions compared with baseline [10]. Therefore, our results in efficient pain processing or compensatory damage in
0 mm −30 mm
ACUP minus OFF status

Deactivation
(a)
−9 mm −20 mm
SHAM minus OFF status

Activation
(b)
−2 mm −24 mm
ACUP minus SHAM

Deactivation
(c)
Figure 7: The fMRI signal increases and decreases in cortical and subcortical brain structures, (1) PAG; (2) pACC, aMCC, and anterior
dmPFC; (3) PHP and HP; (4) PCN, PCC, and RSC; (5) striatum, thalamus, red nucleus, and substantia nigra; (6) lateral temporal cortex; (7)
pMCC; (8) mammillary body.
functionally relevant regions such as the prefrontal cortex and Naqvi’s conclusion that this hemisphere corresponds to the
caudate [24]. Pain is well documented to potentially interrupt affective consequences of pain, whereas the right hemisphere
cognition and sustained attention to a direct action toward corresponds mainly to homeostatic and autonomic control
a painful stimulus or threat [25]. The insula is an important [28].
component of the pain system, and its functions involve
judgment about potential dangers [26]. The results showed 4.2. The Effect of Acupuncture in the Brain Network
higher ReHo values in the right insula, possibly indicating
an increase in the judgment function and evasive actions 4.2.1. Limbic System. Interest in the role of PAG and ACC
during the pain state; because the insula also participates in for pain modulation has a long history [7, 16, 29–32].
learning and memory regarding pain [27], the higher ReHo Anatomically, nociceptive signals can ascend to PAG and
values in the insula indicate increased function. The ACC and the posterolateral thalamus, for which the signals project to
insula exhibited higher ReHo values in the pain matrix during S1, S2, and ACC [21]; moreover, they could directly project
experimental LBP. The negative correlations between ACC through the midline and intralaminar thalamic nuclei to
and the insula were enhanced, suggesting that the anterior other limbic areas, including PAG, ACC, and amygdala [32].
insula reduces the response to peripheral nociceptive stimuli Functionally, investigators reported that PAG demonstrated
via a self-control function. coherence with ACC (rostral and pregenual) in the resting
Furthermore, the brain regions with decreased ReHo val- state and formed a core intrinsic functional ACC-PAG-RVM
ues were concentrated in the left hemisphere, which verifies network for pain modulation [31, 33]. Furthermore, Hui et al.
0 mm −30 mm
ACUP minus OFF status

Deactivation
(a)
−9 mm −20 mm
SHAM minus OFF status

Activation
(b)
Figure 8: The fMRI signal increases evoked by ACUP and SHAM, (1) right insula and frontal operculum cortex; (2) dlPFC; (3) supramarginal
gyrus/angular gyrus; (4) orbitofrontal cortex; (5) lateral temporal cortex and temporal pole.
summarized a series of their studies conducted over the Anatomically, DMN comprises the regions along the ante-
last decade, and found that acupuncture analgesia was rior and posterior midline, the lateral parietal cortex, the
mainly relevant in terms of the deactivations in the limbic- prefrontal cortex, and the medial temporal lobe (MTL); it
paralimbic-neocortical network (LPNN), including PAG and therefore overlaps with the limbic system to a certain degree.
ACC [16]. Consistent with these findings, our results may The precise function of DMN remains debatable; however,
suggest that acupuncture therapy reverses the activation of analysis of its intrinsic activity has revealed that its function
limbic structures evoked by pain, resulting in an analgesic might be divided into the MTL and dmPFC subsystems, with
effect. a midline core (PCC and anterior mPFC) [6].
In 1968, Melzack and Casey described pain in terms Consistent with this finding, ACUP in the ALBP subjects
of its three dimensions: “sensory-discriminative,” “affective- yielded widespread deactivations in the PCC and MTL
motivational,” and “cognitive-evaluative” [34]. Many studies subsystems, including the HP/PHP, pMCC/PCN/PCC/ret-
have proved that the pACC and PAG are involved in not only rosplenial cortex, and the angular gyrus, which play key
acute pain but also emotion [31, 32]. The pACC has been roles in recalling the past or imagination of the future [6,
found to be closely related to the affective network and could 38]. In addition to DMN, the deactivated mammillary body,
be specifically evoked by positive events [35], while PAG has which usually acts as a relay for impulses coming from
been found to be significantly connected with its surrounding the amygdala and HP through the mamillothalamic tract
areas and is important for the control of emotions, for exam- to the thalamus, is part of the larger Papez circuit and is
ple, fear and the affective aspect of pain [31]. Furthermore, involved in storing memory [39]. Furthermore, investigators
the aMCC and mammillary body may also be activated demonstrated that acupuncture could modulate memory
by negative emotion [36, 37]. Based on above analysis, encoding and retrieving in patients with mild cognitive
we speculated that the unpleasantness of acute pain could impairment [40]. Consequently, we propose that acupunc-
induce dysfunction in these emotion-processing regions, and ture reduced spontaneous memory-related cognition, which
acupuncture may be beneficial for treating this dysfunction. might provide psychological relief from pain.
Besides the MTL subsystem, ACUP, also deactivated the
4.2.2. Default Mode Network. The DMN generally shows dmPFC subsystem, including the supramarginal gyrus and
specific spontaneous activations when a person is left undis- the dmPFC. Prior studies have demonstrated that the dmPFC
turbed, for example, lying peacefully in an MRI or positron is linked with lower levels of autonomic outflow regions,
emission tomography scanner. Interestingly, these activations including PAG and the hypothalamus in monkeys and rats,
transform into coordinated deactivations during attention- respectively [41, 42], and with the pACC in humans [43].
demanding tasks such as pain or acupuncture stimuli [5, 6]. Clinical studies further found that acupuncture with the de qi
Table 3: fMRI signal changes evoked by SHAM (SHAM (ON status compared with OFF status)) one-sample 𝑡-test analysis 𝑃 < 0.05, FDR <
0.05.
Peak MNI coordinate
𝑋 𝑌 𝑍
Left insular 13 99 −3.60 −39 −12 15
Left frontal operculum 6 78 −3.46 −40 0 20
Left M1 44 60 −3.01 −23 −10 14
Left SMA 6 271 4.19 −27 3 57
Right dlPFC 46 80 4.01 −48 39 21
Right SMA 6 210 3.69 30 42 24
Left dlPFC 8 189 3.44 37 43 20
Right frontopolar area 9 154 3.23 30 25 30
Left mammillary body 456 5.67 −15 −3 3
Right thalamus 356 4.57 −12 0 0
Right mammillary body 234 3.58 19 23 8
Left thalamus 315 4.43 8 3 3
Amygdala 56 3.01 3 2 7
Left Hp 23 43 4.88 −27 −36 −6
Right PHP 36 32 3.45 −20 −23 0
Orbitofrontal cortex 37 34 3.03 −19 34 3
Right temporal pole 42 35 3.23 −24 −35 0
Left PCN 7 78 4.75 6 −57 36
Right pACC 23 56 3.89 5 −50 42
Left ACC 24 57 3.76 −4 −66 22
Left angular gyrus 19 49 3.35 27 −42 −27
Right angular gyrus 19 54 3.45 −28 −44 −34
Left cerebellum anterior lobe — 46 3.25 34 −22 −20
Right cerebellum anterior lobe — 45 3.24 −32 −10 20
FDR: false discovery rate; MNI: Montreal Neurological Institute; M1: primary motor cortex; SMA: supplementary motor area; dlPFC: dorsolateral prefrontal
cortex; HP: hippocampus; PHP: parahippocampus, PCN: precuneus; pACC: pregenual anterior cingulate cortex; ACC: anterior cingulate cortex.
sensation could inhibit the dmPFC for treating various This view suggests that (1) active cognitive ratings during
psychological problems, such as schizophrenia and anxiety acupuncture evoked stronger de qi sensations than passive
disorders [44, 45]. Moreover, acupuncture could decrease sensory stimuli [47]; (2) stronger de qi sensations evoked by
sympathetic activity and increase parasympathetic activity by acupuncture enhance more cognition than tactile stimulation
inhibiting the dmPFC [46]. [10]; and (3) paying attention to the pain can upregulate pain,
while distraction can downregulate pain [48]. In addition,
4.2.3. Contact between the LPNN and DMN. Regulation of acupuncture stimuli could act as a placebo, and de qi sen-
negative LPNN activity was a notable result of acupuncture. sation ratings may promote this placebo effect [49]. Broadly
This network is thought to have significant relationships with consistent with these views, our result in the ALBP subjects
pain conduction and changes in the brain function network showed that both ACUP and SHAM, along with sensory
involved with acupuncture regulation [16]; DMN of the brain stimuli and cognitive rating, evoke prominent activations in
overlaps with LPNN that is deactivated by acupuncture. dlPFC and pMCC (Figures 7 and 8). Furthermore, the right
Research has shown that, in terms of brain function, DMN insula and frontal operculum cortex activated by ACUP was
interacted with LPNN, with broad activation. We confirmed another important attention-related area [50]. As previously
Fang et al.’s conclusion [11] that this intrinsic organization observed, these four regions were thought to provide a higher
may be a core function of LPNN network in response to level role in attentional control, including continuous mon-
ACUP. itoring of the external world, searching behavior for active
solution derivation, and regulating the skeletomotor system
4.3. Activation Network in Acupuncture Studies. Some in the presence of interfering stimuli [10, 32, 50–52]. Unlike
researchers believe that ACUP serves as a somatosensory- the findings from a previous study [10], we found that ACUP
guided mind-body therapy, which effectively combines evoked stronger de qi sensations in ALBP subjects, inducing
peripheral sensory stimuli and cognitive ratings [10]. weaker activations in both dlPFC and pMCC compared with
Table 4: fMRJ signal changes in the comparison of ACUP minus SHAM (ACUP compared with SHAM) paired 𝑡-test analysis 𝑃 < 0.05, FDR
< 0.05.
Peak MNI coordinate
𝑋 𝑌 𝑍
Left SMA 6 97 −3.75 −24 12 60
Left frontal eye field 8 76 −3.45 −20 10 80
Right insular 13 160 3.79 39 9 9
Right M1 149 3.54 54 10 11
Right frontal eye field 8 54 −3.34 23 −10 70
Right dlPFC 38 42 −3.57 −54 12 −8
Left HP 24 234 −4.46 −12 −24 −6
Left PHP 23 121 −3.89 −10 −22 0
Left mammillary body 23 111 −3.65 −16 −19 22
Left thalamus 56 −2.79 12 34 0
Right PCC 21 76 −4.01 −3 −6 30
Left ACC 45 −2.58 4 5 −45
Right supramarginal gyrus 40 387 −4.34 57 −54 24
Right angular gyrus 22 134 −3.78 60 −34 20
Right precuneus 42 145 −3.89 45 −20 −30
Right thalamus 13 84 −4.09 24 −27 4
Right insular 76 −3.75 23 30 10
Right PHP 35 81 −4.06 24 −24 −18
Right HP 28 45 −3.06 30 −10 −29
Left dmPFC 24 99 −3.78 0 33 18
Left PAG 31 117 −4.31 −9 −48 37
Right cerebellar anterior lobe 234 −4.76 9 −60 40
Left cerebellar anterior lobe 320 −4.32 −18 56 33
Lateral occipital gyrus 10 87 −3.54 2 33 −18
FDR: false discovery rate; MNI: Montreal Neurological Institute; SMA: supplementary motor area; dlPFC: dorsolateral prefrontal cortex; HP: hippocampus;
PHP: parahippocampus; pACC: pregenual anterior cingulate cortex; ACC: anterior cingulate cortex; dmPFC: dorsomedial prefrontal cortex.
SHAM. These differences can be explained by the fact that in the left S1, supplementary motor area, and anterior insula
the de qi sensations were rated at end of each functional compared with noninsertive cutaneous stimulation, although
scan, rather than during each block. Moreover, previous this was not found to be the case in healthy control subjects
researchers found that too little autonomic arousal may fail [55]. Our results in ALBP subjects showed that ACUP in the
to activate the dlPFC, while too much attention focused upon right leg deactivated the left sensorimotor regions (S2, M1,
a task may limit dlPFC function and selection of optimal and insula) while activating the right sensorimotor regions
responses in people with elevated anxiety [50]. Taken (S1, M1, and insula). One reasonable explanation for our
together, we considered that moderate activity in the atten- result is that the acupuncture stimuli may have inhibited the
tion network may be important for acupuncture analgesia. ipsilateral ascending nociceptive inputs and facilitated the
In animal studies, researchers found that the analgesic contralateral inputs to some extent.
effects of manual acupuncture may have mainly resulted from
a C-type afferent, by means of selective blockade of con- 5. Limitation
duction in C- and A𝛿-type afferents [4]. This effect seemed
to act as a diffuse noxious inhibitory control, which also This fMRI-based study of the analgesic mechanism of
mediated C- and A𝛿-type afferents and strongly alleviated acupuncture provides a good foundation for future research.
the initial painful sensation [53]. On the other hand, a However, the study also has some limitations. First, the types
fMRI study showed that moderate-high thermal pain on of data differed, particularly, the baseline and the pain state
the right forearm activated the left sensorimotor regions data, were rs-fMRI type data, whereas the ACUP and SHAM
(S1 and M1), bilateral insula, and S2, while deactivating the data were task fMRI type data. Therefore, we could only
right sensorimotor regions (S1 and M1) [54]. However, a compare the pain state data with the baseline in the ReHo
clinical study in carpal tunnel syndrome subjects showed that model and compare the ACUP or SHAM status data with the
acupuncture in the right hand yielded significant deactivation OFF status data in the GLM model. We could not compare
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Conflict of Interests by puncturing Weizhong (BL 40)’ with blood perfusion imaging
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All authors report no conflict of interests. [13] P. G. Nash, V. G. MacEfield, I. J. Klineberg, S. M. Gustin,
G. M. Murray, and L. A. Henderson, “Bilateral activation of
Authors’ Contribution the trigeminothalamic tract by acute orofacial cutaneous and
muscle pain in humans,” Pain, vol. 151, no. 2, pp. 384–393, 2010.
Yu Shi and Ziping Liu contributed to the work equally and [14] D. Falla, L. Arendt-Nielsen, and D. Farina, “Gender-specific
should be regarded as co-first authors. adaptations of upper trapezius muscle activity to acute nocicep-
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Acknowledgments gence in central processing,” Annual Review of Neuroscience,
vol. 26, pp. 1–30, 2003.
The authors thank Yang JM from the Department of Neu-
[16] K. K. S. Hui, O. Marina, J. Liu, B. R. Rosen, and K. K. Kwong,
rology, Zhujiang Hospital, Southern Medical University in “Acupuncture, the limbic system, and the anticorrelated net-
China for assistance. The authors also thank all subjects for works of the brain,” Autonomic Neuroscience: Basic & Clinical,
the assistance in the scanning. This work was supported vol. 157, no. 1-2, pp. 81–90, 2010.
by Grants from the National Natural Science Foundation [17] R. P. Dhond, C. Yeh, K. Park, N. Kettner, and V. Napadow,
of China (NNSFC), China (Contract Grant no. 81473769) “Acupuncture modulates resting state connectivity in default
and the Natural Science Foundation of Guangdong Province, and sensorimotor brain networks,” Pain, vol. 136, no. 3, pp. 407–
China, Contract Grant no. 2014A030313335. 418, 2008.
[18] Y. Zang, T. Jiang, Y. Lu, Y. He, and L. Tian, “Regional homogene-
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http://dx.doi.org/10.1155/2015/641286
Research Article
Effect of Repeated Electroacupuncture
Intervention on Hippocampal ERK and p38MAPK
Signaling in Neuropathic Pain Rats
Jun-ying Wang, Shu-ping Chen, Yong-hui Gao, Li-na Qiao,

Jian-liang Zhang, and Jun-ling Liu
Department of Physiology, Institute of Acupuncture and Moxibustion, China Academy of Chinese Medical Sciences,
Correspondence should be addressed to Jun-ling Liu; junling liu@aliyun.com
Received 24 November 2014; Revised 7 April 2015; Accepted 7 April 2015
Copyright © 2015 Jun-ying Wang et al. This is an open access article distributed under the Creative Commons Attribution License,
Results of our past studies showed that hippocampal muscarinic acetylcholine receptor (mAChR)-1 mRNA and differentially
expressed proteins participating in MAPK signaling were involved in electroacupuncture (EA) induced cumulative analgesia in
neuropathic pain rats, but the underlying intracellular mechanism remains unknown. The present study was designed to observe
the effect of EA stimulation (EAS) on hippocampal extracellular signal-regulated kinases (ERK) and p38 MAPK signaling in rats
with chronic constrictive injury (CCI) of the sciatic nerve, so as to reveal its related intracellular targets in pain relief. After CCI, the
thermal pain thresholds of the affected hind were significantly decreased compared with the control group (𝑃 < 0.05). Following
one and two weeks’ EAS of ST 36-GB34, the pain thresholds were significantly upregulated (𝑃 < 0.05), and the effect of EA2W was
remarkably superior to that of EA2D and EA1W (𝑃 < 0.05). Correspondingly, CCI-induced decreased expression levels of Ras, c-
Raf, ERK1 and p-ERK1/2 proteins, and p38 MAPK mRNA and p-p38MAPK protein in the hippocampus tissues were reversed
by EA2W (𝑃 < 0.05). The above mentioned results indicated that EA2W induced cumulative analgesic effect may be closely
associated with its function in removing neuropathic pain induced suppression of intracellular ERK and p38MAPK signaling in
the hippocampus.
1. Introduction in long-term potentiation (LTP) [6], impaired enriched-

environment neurogenesis [9], and altered synaptic plasticity
It has been well-documented that patients with chronic [10]. Increasing evidence has demonstrated the involve-
pain often experience sustained chronic psychological and ment of hippocampus in acupuncture analgesia [11–13] and
physical stress and exhibit increased anxiety, depression, and acupuncture signal processing [14, 15].
deficits in working memory [1–3]. Results of a pilot study Our experimental studies demonstrated that in chronic
showed that in elderly patients with chronic pain, a reduced constrictive injury- (CCI-) induced neuropathic pain rats,
hippocampal volume and lower levels of hippocampal N- the resultant cumulative analgesic effect of repeated electroa-
acetylaspartate to creatine ratios (NAA/Cr) were found [4]. cupuncture stimulation (EAS) of Zusanli (ST36)-Yangling-
The hippocampus, an important region of the limbic system, quan (GB34) is closely associated with its effects in upregu-
has been shown to be complicated in pain processing, lating the decreased hippocampal synaptophysin immunoac-
particularly under chronic pain conditions [1–3]. tivity [16], muscarinic acetylcholine receptor (mAChR)-1
Animal studies showed hippocampal abnormalities in mRNA and choline acetyl transferase (ChAT) mRNA expres-
animal models of chronic pain including short-term working sion [17], and improving synaptic plasticity of nerve cells in
memory dysfunction [5], recognition memory deficits [6], the hippocampal CA3 region shown by electron transmis-
abnormal cytokine (IL-1𝛽 mRNA) expression [7, 8], deficits sion microscope [18]. Differential proteomics analysis and
Western blotting validation indicated that the 19 hippocampal 2.3. Chronic Neuropathic Pain. The chronic pain model was
differentially-depressed proteins involving repeated EAS- established by ligature of the left sciatic nerve with reference
induced pain relief are those participating in metabolic, to modified Bennett’s and Xie’s methods [27]. Under anes-
physiological, and cellular processes, and so forth, and one thesia (with mixture solution of urethane 28 mg/100 g plus
of the top three canonical pathways identified is “mitogen- chloralose (Sigma, 3.3 mg/100 g)) and routine sterilization,
activated protein kinase (MAPK) signaling” [19]. the left sciatic nerve was exposed at the midthigh level by
MAPK is an important protein molecule for intracellular blunt dissection through the biceps femoris muscle. Four
signal transduction and is involved in many physiological constrictive ligatures (4–0 surgical suture) were tied around
and pathological processes of biological activity. The MAPK the nerve at the distal end close to the bifid site at spaces
family mainly includes extracellular signal-regulated kinases of about 1.0 mm apart. The ligature was alright till the
(ERK), p38 MAPK, and c-Jun N-terminal kinase/stress- local moderate muscular contraction of the leg could be
activated protein kinase (JNK/SAPK), which represent three seen clearly. After local application of antibiotic (sodium
separate signaling pathways [20, 21]. The MAPKs signaling penicillin, 9,000–10,000 U/rat), the muscle and skin were
cascades from extracellular stimuli into a variety of intra- sutured in layers. For rats of the control group, the left
cellular responses are involved in various cellular functions sciatic nerve was just exposed without ligature. For reducing
experimental variability, all the operations were finished by
by sequential activation of MAPKKK, MAPKK, MAPK, and
the same one operator.
transcription factors [22]. ERK signaling pathway is a cascade
involving sequential activation of Ras, Raf, mitogen-activated
protein kinase (MEK), ERK, p38MAPK, MKK3, 6, and p38 2.4. Electroacupuncture Treatment. Bilateral “Zusanli” (ST36)
(𝛼, 𝛽, 𝛾, 𝛿) [20]. By using MAPK inhibitors targeting ERK, and “Yanglingquan” (GB34) were punctured with stainless-
p38 MAPK, and JNK in combination with LTP recording steel acupuncture needles (Gauge 28, 0.20 mm in diameter)
in the dorsal hippocampus formation (HF), Liu et al. [23] to a depth of about 4 mm, respectively, and stimulated
demonstrated that the specific members of the MAPK family electrically by using a HANS EA Stimulator (LH202, made
in China). EA (2/15 Hz, 1 mA) was given to rats for 30 min,
might mediate pain-associated spatial and temporal plasticity
once a day, continuously for one week (from day 12 on after
in the HF. In addition, it has been shown that after periph-
CCI), 2 weeks (from day 4 on after CCI), and 2 days (from
eral nerve injury, ERK and p38 MAPK were activated and
day16 on after CCI), respectively.
increased in their expression levels in the spinal dorsal horns
[24–26]. However, there has been no any research on the
effect of repeated EAS on changes of ERK signaling pathway 2.5. Thermal Pain Threshold Detection. When thermal hyper-
and p38 MAPK in the hippocampus in neuropathic pain algesia test was conducted, the animal was put into a black
animals. For this reason, the present study was designed to cloth bag with the hindlimbs and tail exposed to move freely.
investigate the relationship between EAS-induced cumulative A mobile radiant heat source (a high-intensity light beam
analgesia and activities of ERK and p38MAPK signaling in of radiant heat dolorimeter) was focused onto the plantar
the hippocampus in CCI rats for revealing the underlying surface of the hindpaw. The paw withdrawal latency (PWL)
intracellular mechanism of EAS analgesic target. (i.e., pain threshold, PT) of the rat’s bilateral footplates was
detected 3 times, with an interval of about 5 min between
two detections. In order to avoid potential tissue damage, the
2. Materials and Methods cutoff time of the radiant heat radiation was set at 20 sec. The
2.1. Ethic Statement. The protocols of the present study were mean PT before CCI operation was used as the control value,
and 4 days after CCI operation, PT was detected again. For
approved by the Institute of Acupuncture and Moxibustion,
rats of CCI + EA groups, PT was detected on the following
China Academy of Chinese Medical Sciences. The study was
day for observing the posteffect of EA. In order to minimize
carried out in accordance with the recommendation in the
the animal individual difference, the difference value of PWL
Guidelines for Declaration of the National Institutes of Health between the healthy and the affected footplates was used as
Guide for Care and Use of Laboratory Animals (publication the pain score.
number 80-23, revised 1996). All surgical operations were
performed under anesthesia, and all efforts were made to
minimize animals’ sufferings. 2.6. Western Blot. The right hippocampus was taken to be
frozen in liquid nitrogen and stored at −80∘ C until use. Total
protein was extracted first from the tissue in RIPA Lysis Buffer
2.2. Animals and Grouping. Adult male Wistar rats (200– containing protease and phosphatase inhibitors (Roche) by
250 g), purchased from Beijing Union Medical College, were using a tissue homogenizer. The tissue lysate was then cen-
acclimatized to standard laboratory conditions (about 12 h trifuged at 13000 rpm at 4∘ C for 20 min, and protein concen-
alternate light-dark cycle) of our institute’s environment first tration of supernatants was determined using a bicinchoninic
for a week and were given free access to standard chow acid (BCA) protein assay kit (Thermo Scientific). Equivalent
pellet diet and water. The rats were randomly assigned to amount of protein (50 𝜇g/tissue lysate) in each sample was
5 groups: control, model (chronic constrictive injury, CCI), loaded per lane and separated by 5% or 8% sodium dodecyl
CCI + EA2D (days), CCI + EA1W (week), and CCI + EA2W, sulfate polyacrylamide gel electrophoresis (SDS-PAGE) for
with 14 rats in each group. about 60 min at 90/160 V and then electrotransferred onto
polyvinylidene difluoride (PVDF) membrane for 150 min at 7

90 m A. The membranes were blocked with 5% bovine serum 6 *
albumin (BSA, Amresco, USA) solution for 30 min at room
temperature. The membranes were incubated with primary 5 *
antibody Ras protein (1 : 5000, Cell Signaling Technology), ☆△◇
The pain score

4
c-Raf protein (1 : 2000, Cell Signaling Technology), MEK1 ☆△
protein (1 : 10000, abcam), P44/42 (1 : 10000, Cell Signaling 3
Technology), P-P44/42 (1 : 5000, Cell Signaling Technology), 2 ☆△◇
P38 (1 : 1000, abcam), and P-P38 (1 : 2000, Epitomics) at 4∘ C
1
overnight. After washing, the membranes were incubated
with secondary antibody (1 : 20000 diluted with goat anti- 0
rabbit Immunoglobulin (Ig) G or 1 : 10000 diluted with Before 4th day after 8th day after 16th day after
−1 operation operation operation operation
goat anti-mouse IgG) conjugated to horseradish peroxidase
(Jackson Immuno Research Laboratories) for 1 h at room Normal control CCI + EA1W
temperature on the following day. The membranes were CCI CCI + EA2W
developed using an enhanced chemiluminescence (ECL) CCI + EA2D
detection system to transfer to film. For densitometric Figure 1: Effect of EA of ST36-GB34 on pain scores in CCI rats of
analyses, the blots were scanned and quantified using Total different groups. Thermal pain thresholds after injury and EA are
Lab Quant analysis software (TotalLab Limited, England), presented as mean ± SD (𝑛 = 11 in each group; ∗ 𝑃 < 0.05 compared
and the result was expressed as the ratio of target gene with the sham control group; f 𝑃 < 0.05, compared with the CCI
immunoreactivity to 𝛽-action immunoreactivity. group; △ 𝑃 < 0.05, compared with the CCI + EA2D group; ⬦ 𝑃 <
0.05, compared with the CCI + EA1W group). Pain score = the paw
2.7. RNA Isolation and Quantitative Real-Time PCR. The right withdrawal latency (PWL) of the healthy side (right) – PWL of the
affected side (left).
hippocampus samples were excised and ground into powder
in liquid nitrogen. Total RNA was isolated from hippocampus
with Trizol (CW0581, CWbio. Co. Ltd., Beijing, China)
and then reversely transcribed using a cDNA Synthesis Kit (mean ± SD) and analyzed by two-way repeated measures
(CW0744, CWbio. Co. Ltd., Beijing, China). The reverse- ANOVA, followed by post hoc test for least significant
transcribed products were amplified. The primer sequences difference (LSD) to determine differences between every two
used were as follows: ERK1: forward: 5󸀠 -CGTTCAGATGTC- groups. Statistical significance was accepted with 𝑃 < 0.05.
GGTGTC-3󸀠 , reverse: 5󸀠 -AAAGGAGTCAAGAGTGGG-
3󸀠 ; ERK2: forward: 5󸀠 -CCAGAGTGGCTATCAAGAAG-3󸀠 , 3. Results
reverse: 5󸀠 -GGATGTCTCGGATGCCTA-3󸀠 ; p38 MAPK:
forward: 5󸀠 -GTACCTGGTGACCCATCTC-3󸀠 , reverse: 5󸀠 - 3.1. Effect of EA on Pain Response after CCI. The pain score
GATTATGTCAGCCGAGTGTAT-3󸀠 ; 𝛽-actin: forward: 5󸀠 - is referred to the paw withdrawal latency of the difference
GGAGATTACTGCCCTGGCTCCTA-3󸀠 , reverse: 5󸀠 -GAC- between the healthy and the surgical footplates in the present
TCATCGTACTCCTGCTTGCTG-3󸀠 . Quantitative real- paper. Results (Figure 1) indicated that before CCI, the pain
time- (QRT-) PCR was performed in 96-well plates using the scores of the control (sham operation), CCI model, CCI +
QRT-PCR detection systems (AB7500, Applied Biosystems, EA2D, CCI + EA1W, and CCI + EA2W groups had no
USA). Three different biological replicates for each sample significant difference (𝑃 > 0.05). After CCI, the pain scores
were performed. All the cDNA samples were amplified in of the CCI group were evidently higher than those of the
triplicate from the same RNA preparation and the mean value control group (𝑃 < 0.05), suggesting a hyperalgesia after CCI.
was calculated. Each reaction included 2 𝜇L of cDNA, 10 𝜇L On day 4 after CCI, the pain scores of the model group and
of REALSYBR Mixture (2x), 0.8 𝜇L (10 𝜇mol/𝜇L) of both those of the CCI + EA2D and CCI + EA1W and CCI + EA2W
forward and reverse primers, and 7.2 𝜇L of PCR-grade water, groups were comparable (𝑃 > 0.05), while on day 8, the pain
equating to a final volume of 20 𝜇L. PCR was performed scores of the CCI + EA2W group and, on day 20, those of
under following conditions: 10 min at 95∘ C, followed by 40 the CCI+EA1W and CCI + EA2W were obviously lower than
cycles of 15 s at 95∘ C, and 60 s at 60∘ C. Then, the fluores- those of the model group (𝑃 < 0.05), and the effect of the
cence acquisition after each cycle was performed. Finally, a CCI + EA2W group was significantly better than that of the
dissociation curve was generated by increasing temperature CCI + EA2D and CCI + EA1W groups (𝑃 < 0.05), suggesting
from 65∘ C to 95∘ C in order to verify primer specificity. All a cumulative analgesic effect of repeated EAS of ST36-GB34.
samples for each reference gene were run on the same plate
to avoid between-ran variations. The relative expression was 3.2. Effect of EA on Expression of Hippocampal Ras and C-
calculated in accordance with the ΔΔCT method. Relative Raf Protein in Different Groups. Ras is a membrane-associ-
mRNA levels were expressed as 2−ΔΔCT values. ated guanine nucleotide-binding protein that is normally
activated in response to the binding of extracellular sig-
2.8. Statistical Analysis. The data collected in the present nals [28], and the Raf kinase mediates the transduction
study were presented as mean ± standard deviation of proliferative and differentiative signals from a variety of
Ras c-Raf
GAPDH GAPDH
1.6 1 2 3 4 5 1.4 1 2 3 4 5 ☆
1.4 ☆△ 1.2
1.2
1 *
Relative intensity of c-Raf

Relative intensity of Ras
1 *
0.8
0.8
0.6
0.6
0.4
0.4
0.2 0.2
0 0
Normal CCI CCI + CCI + CCI + Normal CCI CCI + CCI + CCI +
control EA2D EA1W EA2W control EA2D EA1W EA2W
(a) (b)
Figure 2: Effect of EAS on expression levels of hippocampal Ras and c-Raf proteins in different groups. After EA treatment, hippocampus
tissues were prepared for assaying expression levels of Ras, c-Raf, and other related kinases (MEK, ERK, p38 MAPK) by Western blot. Data
are presented as mean ± SD (∗ 𝑃 < 0.05, compared with the sham control group; f 𝑃 < 0.05, compared with the CCI group; △ 𝑃 < 0.05,
compared with the CCI + EA2D group; 𝑛 = 5 in each group). (a) Top panel shows immunoblots of Ras and c-Raf proteins and GAPDH in
different groups: (1) sham control group, (2) CCI group, (3) CCI + EA2D group, (4) CCI + EA1W group, and (5) CCI + EA2W group. GAPDH:
glyceraldehyde-3-phosphate dehydrogenase (housekeeping gene); (b) histograms show the relative expression of Ras and c-Raf proteins in
the 5 groups.
cell surface receptors to the nucleus and is the entry point 3.4. Effect of EA on Hippocampal ERK and p-ERK mRNA and
to the MAPK/ERK-1/2 signaling pathway, which controls Protein Expression. Like MEK, ERK exists in two isoforms
fundamental cellular functions [29]. (1 and 2). In order to identify changes of hippocampal
Following CCI, hippocampal Ras and c-Raf protein ERK1/2 in both mRNA and protein expression levels, we
expression levels were significantly downregulated in com- conducted real-time PCR and Western blot measurements.
parison with those of the control group (𝑃 < 0.05, Figures Compared with the control group, the expression levels of
2(a) and 2(b)). After EAS of ST36 and GB34, both Ras and hippocampal ERK1/2mRNA and ERK1/2 protein in the CCI
c-Raf expression levels were considerably upregulated only group had no significant changes (𝑃 > 0.05), except for
in the CCI + EA2W group (𝑃 < 0.05), rather than in the a marked upregulation of ERK1 protein expression in the
CCI + EA2D and CCI + EA1W groups (𝑃 > 0.05) in spite CCI + EA2W group in comparison with the CCI group
of mild upregulation. The effect of the CCI + EA2W group (𝑃 < 0.05, Figures 4(a) and 4(b)). Further tests revealed that
in upregulating Ras protein was significantly better than that the relative expression of p-ERK1/2 protein was considerably
of the CCI + EA2D and CCI + EA1W groups (𝑃 < 0.05). downregulated in the CCI group compared with the control
No significant difference was found between the CCI + EA2D group (𝑃 < 0.05, Figure 4(c)) and obviously upregulated in
and CCI + EA1W groups (𝑃 > 0.05). the CCI + EA2D, CCI + EA1W, and CCI + EA2W groups after
EAS (𝑃 < 0.05). There was no significant difference among
3.3. Effect of EA on Hippocampal MEK and p-MEK1/2 Protein the three EAS groups in hippocampal p-ERK1/2 protein
Expression in Different Groups. MEK1/2 (MKK1/2) are the expression levels (𝑃 > 0.05, Figure 4(c)).
upstream kinases of ERK signaling. Compared with the
control group, the expression levels of hippocampal MEK and 3.5. Effect of EA on Hippocampal p38 MAPK mRNA and Pro-
p-MEK1 proteins had no significant changes in the CCI, CCI tein Expression. Activation of MAPK is the final step of
+ EA2D, CCI + EA1W, and CCI + EA2W groups (𝑃 > 0.05, intracellular phosphorylation cascade reactions in response
Figure 3(a)), while that of p-MEK2 protein was significantly to extracellular signal. Compared with the control group,
downregulated after CCI (𝑃 < 0.05, Figure 3(b)). Following hippocampal p38MAPKmRNA and p-P38MAPK protein
EAS of ST36-GB34, p-MEK2 expression had a slight upregu- expressions were significantly and moderately downreg-
lation in the three EAS groups (𝑃 > 0.05) without significant ulated, respectively, in the CCI group (𝑃 < 0.05,
differences among the three groups (𝑃 > 0.05). Figures 5(a) and 5(c)). Following EA of ST36-GB34, both
p-MEK1/2
GAPDH
1 2 3 4 5
1.6
MEK
1.4
GAPDH
1 2 3 4 5 1.2
Relative intensity of p-MEK1/2

1.2 1 *
1
Relative intensity of MEK
0.8
0.8
0.6
0.6
0.4
0.4
0.2
0.2
0 0
Normal CCI CCI + CCI + CCI + p-MEK1 p-MEK2
control EA2D EA1W EA2W
Normal control CCI + EA1W
CCI CCI + EA2W
CCI + EA2D
(a) (b)
Figure 3: Effect of EA on expression levels of hippocampal MEK, p-MEK proteins in different groups. Data are presented as mean ± SD
(∗ 𝑃 < 0.05, compared with the sham control group; 𝑛 = 5 in each group). (a) Upper panel shows representative immunoblots of MEK protein
in the 5 groups: (1) sham control group, (2) CCI group, (3) CCI + EA2D group, (4) CCI + EA1W group, and (5) CCI + EA2W group; lower
histograms show the relative expression levels of MEK protein in the 5 groups. (b) The lower histograms show the relative expression levels
of p-MEK1 and p-MEK2 proteins in the five groups; upper panel shows the representative immunoblots of MEK1/2 proteins and GAPDH in
different groups.
p38MAPKmRNA and p-P38MAPK protein were obviously p-ERK1/2 protein, and p38MAPK mRNA were obviously
upregulated only in the CCI + EA2W group (𝑃 < 0.05). downregulated and that of p-p38MAPK protein was mod-
There were no significant changes of hippocampal p38MAPK erately downregulated in spite of the fact that there was
protein expression in the five groups and p38MAPK mRNA no statistical significance. It suggests an inhibition of hip-
and p-p38MAPK protein expression in the CCI + EA2D and pocampal ERK/MAPK signaling after CCI in neuropathic
CCI + EA1W groups (𝑃 > 0.05, Figures 5(a), 5(b), and 5(c)). pain rats. These results of hippocampal molecules are also
basically identical to those of Mutso et al. report [10] which
showed reduced ERK expression and phosphorylation in the
4. Discussion hippocampus in spared nerve injury (SNI) (tight ligature and
severing of the tibial and common peroneal nerves) mice
Results of the present study showed that following CCI, the and to Liu and colleagues’ study [23] about an involvement
pain threshold of the affected paw was significantly lowered of ERK and p38MAPK in pain processing in the dorsal
and the difference values of PWL of the bilateral paws hippocampus formation, in which ERK and p38 MAPK
(pain scores) were apparently increased, peaking on day 8 seemed to play opposing roles, with the former positively
after CCI, which is similar to Bennett’s and Xie’s outcomes involved and the latter negatively involved. CCI may be
[27]. Following EAS of ST36-GB34, the pain threshold was considered to be chronic stress stimulation and chronic pain
markedly increased in both EA1W and EA2W groups, but not often resulting in depression. Thus, some molecular changes
in the EA2D group, presenting a cumulated analgesic effect of the hippocampus under chronic stress and depression
after repeated EAS, which are identical to our results of past conditions may also be used as references. It was reported
studies [17, 30, 31] and related reports [32, 33]. that chronic stress exposure caused a reduction in p-ERK and
Correspondingly, after CCI, the expression levels of p-CREB expression in the hippocampus of rats [34, 35]. In
intracellular Ras, c-Raf, p-MEK proteins, ERK2 mRNA, terms of depression caused by chronic pain [1, 10, 36–38],
Relative expression of ERK1 mRNA
0.7
1 2 3 4 5
0.6 ERK1/2
0.5
1.4 GAPDH
0.4
☆
0.3
1.2
0.2
0.1
1
Relative intensity of ERK1/2

0
Normal CCI CCI + CCI + CCI +
0.8
Relative expression of ERK2 mRNA
20
18 0.6
16
14
12 0.4
10
8
6 0.2
4
2
0 0
Normal CCI CCI + CCI + CCI + ERK1 ERK2
Normal control CCI + EA1W
CCI CCI + EA2W
CCI + EA2D
(a) (b)
p-ERK1/2
GAPDH
1 2 3 4 5
1.2
Relative intensity of p-ERK1/2
1
☆
0.8 ☆
☆
0.6
*
0.4
0.2
0
(c)
Figure 4: Effect of EA on expression levels of hippocampal ERK1/2mRNA and ERK1/2 and p- ERK1/2 protein in different groups.
Hippocampal ERK1/2mRNA expression levels were assessed by real-time PCR and ERK1/2 protein expressions were detected by Western
blot. Data are presented as mean ± SD (∗ 𝑃 < 0.05, compared with the sham control group; f 𝑃 < 0.05, compared with the CCI group; 𝑛 = 6
in each group for real-time PCR; 𝑛 = 5 for each group for western blot); (a) histograms show the expression levels of ERK1/2 mRNA. (b)
The top panel shows the representative immunoblots of ERK1/2 proteins in the 5 groups: (1) sham control group, (2) CCI group, (3) CCI +
EA2D group, (4) CCI + EA1W group, and (5) CCI + EA2W group. The histograms show relative expression levels of ERK1/2 proteins in the
5 groups. (c) The upper panel shows the representative immunoblots of p-ERK1/2 proteins in the 5 groups. The lower bar graph shows the
relative expression of p-ERK1/2 proteins in the 5 groups.
p38MAPK
GAPDH
1 2 3 4 5
0.7 1.4
Relative expression of p38MAPK mRNA
Relative intensity of p38MAPK

0.6 1.2
0.5 1
0.4 0.8
☆△
0.3 0.6
0.2 0.4
0.1 0.2
*
0 0
Normal CCI CCI + CCI + CCI + Normal CCI CCI + CCI + CCI +
control EA2D EA1W EA2W control EA2D EA1W EA2W
(a) (b)
p-p38MAPK
GAPDH
1 2 3 4 5
1.2
☆
Relative intensity of p-p38MAPK
1.15
1.1
1.05
1
0.95
0.9
0.85
0.8
(c)
Figure 5: Effect of EA on expression levels of hippocampal p38MAPKmRNA and p38MAPK and p-p38MAPK proteins in different groups.
Hippocampal p38MAPK mRNA and p-38MAPK and p-p38MAPK protein expression levels were assayed by real-time PCR and Western
blot, respectively. Data are presented as mean ± SD (∗ 𝑃 < 0.05, compared with the sham control group; f 𝑃 < 0.05, compared with the CCI
group; △ 𝑃 < 0.05, compared with the CCI + EA2D group; 𝑛 = 6 in each group for real-time PCR; 𝑛 = 5 for each group for Western blot).
(a) Histograms of real-time PCR show the expression levels of p38MAPKmRNA in the 5 groups; (b) the top panel shows the representative
immunoblots of p38MAPK protein and GAPDH in (1) sham control group, (2) CCI group, (3) CCI + EA2D group, (4) CCI + EA1W group,
and (5) CCI + EA2W group. The histograms show the relative expression levels of p38MAPK protein in the 5 groups. (c) Upper panel shows
the representative immunoblots of p-p38MAPK in (1) normal control group, (2) CCI group, (3) CCI + EA2D group, (4) CCI + EA1W group,
and (5) CCI + EA2W group. The histograms show the relative expression levels of p-p38MAPK protein in the 5 groups.
it was demonstrated that chronic unpredictable stress (CUS) injection mimicking transient or persistent pain equally ini-
suppressed p-ERK, p-ERK1/2, and p-CREB expression in the tiated an intense and long-lasting activation of hippocampal
hippocampus. ERKs and ERK1 which were more remarkably activated than
On the other hand, controversial results do exist; for ERK2 in the hippocampus. The possible explanations for
instance, it was reported that 14 days of stress induced an the discrepancy may lie in the difference in stress category,
increase in p-ERK1/2 and p-CREB expression in the hippo- duration, and other experimental procedures, and the acute
campus in rats with infraorbital nerve injury [39]. Under pain is quite different from chronic pain in the underlying
acute conditions, Guo et al. [22] observed that in the hip- mechanisms. Moreover, in the dorsal horns of the spinal cord,
pocampus of naı̈ve rats, intraplantar saline or bee venom the ERK signaling pathway plays an important role in the
genesis and maintenance of pain, which exhibited upregu- our past partial research results, a complete network linking
lation of the expression of ERK and phosphorylated ERK the presynaptic synaptophysin, mAChR, and Ras-Raf-MEK-
proteins under peripheral nerve and tissue injury conditions ERK pathway and synaptic remodeling [18] may participate in
[27, 40, 41]. the cumulative analgesic effect of EAS in neuropathic pain.
Just as those mentioned above, p38 MAPK, an impor- There have been no similar reports available about the
tant member of the MAPKs, plays an important role in effect of EAS on hippocampal ERK and MAPK signaling
the development of central sensitization in responding to in neuropathic pain animal models up to now. Therefore,
chronic nociceptive stimulation shown at the spinal cord we have no way to compare our outcomes with others’
level. Following peripheral nerve injury, p38MAPK and ERK outcomes. However, some results may be used as a reference.
were activated in spinal microglia, and JNK was activated
For example, in depression model rats, EA could reverse CUS
in astrocytes [24, 42]. However, in the hippocampus, there
induced considerable upregulation of p-ERK expression,
has been no direct evidence for its involvement in pain
ratio of p-ERK1/2 to ERK1/2 and the ratio of p-CREB to
processing. In view of chronic neuropathic pain induced
complications as persistent stress, depression, deficits of CREB in the hippocampus [47], or enhanced the activation
memory, and abnormal neural plasticity changes, some find- of hippocampal ERK signaling pathway [48], suggesting an
ings may be used as reference evidence supporting our results. involvement of hippocampal ERK–CREB signaling in EAS-
For example, as a mediator of cellular stresses, p38MAPK was induced antidepressant-like effects. At the spinal level, EAS
implicated in depression induced by forced swim tests and could suppress complete Freund’s adjuvant- (CFA-) induced
tail suspension tests, exhibiting an intensive phosphorylation activation or phosphorylation of p38MAPK in rats with
of PKC-dependent ERK1, ERK2, JNK, and p38MAPK in inflammatory pain [49, 50]. In contusion injury induced
the hippocampus [43]. However, in CUS rats with impaired below-level neuropathic pain rats, acupuncture stimula-
spatial memory, significantly decreased p-CREB and pJNK tion of Shuigou (GV26) and Yanglingquan (GB34) relieved
levels, but without statistical changes in CREB, ERK1/2, mechanical allodynia and thermal hyperalgesia and simul-
p-ERK1/2, p38MAPK, p-P38MAPK, and JNK levels, were taneously inhibited neuropathic pain induced activation of
found in the hippocampus [44]. p38MAPK and ERK in microglia at the L4-5 spinal cord.
Regarding the effect of EAS of bilateral ST36-GB34 on Injection of p38MAPK or ERK inhibitors attenuated neuro-
hippocampal ERK and p38MAPK signaling in the present pathic pain [51]. These results denote that intracellular ERK
study, following two weeks’ EAS, along with the appearance of and p-38 MAPK signaling pathways in the central nervous
cumulative analgesia, the CCI-induced decreased expression system are involved in nociceptive information processing in
levels of Ras, c-Raf, ERK1, p-ERK1/2 proteins, and p38 chronic pain model animals.
MAPK mRNA and p-pMAPK protein were considerably
and gradually upregulated in the hippocampus, denoting a
normalizing trend of functional activities of nerve cells under
EAS-induced pain relief conditions. Most of those proteins 5. Conclusion
were upregulated but had no significant changes after 2 days In conclusion, results of the present study once again demon-
and one week’s EAS, suggesting that two weeks’ EAS has a strated the cumulative analgesic effect of repeated EAS of
cumulative effect in upregulating the activities of ERK and p- ST36-GB34 in CCI-induced neuropathic pain rats and reduce
38 MAPK signaling along with the appearance of cumulative CCI-induced downregulation of Ras, c-Raf, ERK1, p-ERK1/2
analgesic effect. These results are also consistent with our past proteins, and p38 MAPK mRNA and p-pMAPK protein in
results about expression levels of cellular membrane receptors the hippocampus, suggesting an involvement of both ERK
including mAChR1 mRNA and protein [17] and presynaptic and p38 MAPK signaling of hippocampal nerve cells in EAS-
synaptophysin [16] in which two weeks’ EAS evidently sup- induced pain relief. It is sure that this conclusion should be
pressed CCI-induced decrease of their expression in CCI rats. further confirmed by other approaches in the future.
These results indicate that the EAS targets multiple signal
transmission sites from extracellular to intracellular events
during cumulative analgesia induction, and intracellular ERK
and p-38 MAPK signal pathways play an important role in Conflict of Interests
this pain processing. As we know that mAChRs are attributed
to G protein-coupled receptors (GPCRs) which are critical None of the authors has any other conflict of interests related
players in converting extracellular stimuli into intracellular to this paper.
signals in response to various signaling inputs, and these sig-
nal inputs have to be integrated for the processing of complex
biological responses. Chan et al. proved that G protein signals Acknowledgments
can be integrated at the level of MAPK, resulting in differ-
ential effects on ERK, JNK, and p38 MAPK in human brain The present study was supported by National Natural Science
neuroepithelioma cells as a neuronal model [45]. Despite a Foundation of the People’s Republic of China (30472241,
great variety of components of the MAPK/ERK signaling 81202762, key Project: 90709031) and the Ministry of Science
cascade, the architecture of the signal pathway is usually and Technology of PRC (“973” Projects 2007CB512505 and
known as the Ras-Raf-MEK-ERK pathway [46]. Combining 2013CB531904).
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http://dx.doi.org/10.1155/2015/871327
Review Article
Eye Acupuncture Treatment for Stroke: A Systematic
Review and Meta-Analysis
Zeng-Hua Bai,1 Zhi-Xing Zhang,1 Chun-Ri Li,1 Mei Wang,1 Meong-Ju Kim,2 Hui Guo,3,4
Chun-Yan Wang,3,4 Tong-Wu Xiao,5 Yuan Chi,1 Lu Ren,1 Zhong-Yue Gu,1 and Ran Xu6
1
Liaoning University of Traditional Chinese Medicine, 79 Chongshan East Road, Huanggu District, Shenyang 110847, China
2
Department of Alternative Medicine, Nambu University, Kwangju 506-824, Republic of Korea
3
Department of Gynecologic Oncology, Liaoning Cancer Hospital & Institute, Shenyang 110042, China
4
Dalian Medical University, Dalian 116044, China
5
Yangxin People Hospital, Binzhou 251800, China
6
Benxi City Hospital for Infectious Diseases, Benxi 117022, China
Correspondence should be addressed to Zhong-Yue Gu; 470199@163.com and Ran Xu; benxixuran@sina.com
Received 17 December 2014; Revised 2 May 2015; Accepted 5 May 2015
Copyright © 2015 Zeng-Hua Bai et al. This is an open access article distributed under the Creative Commons Attribution License,
There were applications of eye acupuncture for stroke patients. Unfortunately, similar to many other Traditional Chinese Medicine
(TCM) treatments, it lacks comprehensive evaluation and system review for its effect and safety. Objective. This study is a systematic
review to appraise the safety and effectiveness of eye acupuncture for stroke. Methods. “Eye acupuncture therapy” in eleven databases
was searched by randomized controlled trials and quasi-randomized controlled trials. The search activity was ended in April 2014.
The data were extracted and assessed by three independent authors. Rev Man 5.0 software was used for data analysis with effect
estimate presented as relative risk (RR) and mean difference (MD) with a 95% confidence interval. Results. Sixteen trials (1120
patients) were involved with generally poor methodological quality. The study indicated that when eye acupuncture was combined
with western medicine compared to western medicine, there was a significant difference in the areas of mental state, swallow
function, and NDS. When eye acupuncture was combined with western medicine and rehabilitation compared to western medicine
and rehabilitation, there was significant difference in the changes of SSS, FMA, and constipation symptoms evaluation. No adverse
events or side effects have been reported. Conclusions. The current evidence is insufficient and the rigorously designed trials are
warranted.
1. Introduction professor of Liao Ning University of Traditional Chinese

Medicine, in the early 1970s.
Stroke is a neurological deficit that attributed to an acute focal The idea of eye acupuncture was inspired by TCM
central nervous system damage caused by vascular problems,
theory. In his eye acupuncture theory, for the purpose of
such as cerebral infarction, intracerebral hemorrhage, and
both diagnosis and treatment of disease, Dr. Peng divided
subarachnoid hemorrhage. It is a major cause of disability and
the eye into four regions, eight areas, and thirteen points
death worldwide [1]. The burden of ischaemic and haemor-
rhagic stroke have increased between 1990 and 2010 in terms [3]. Eye acupuncture therapy is thought to be a kind of
of the absolute number of people with incident ischaemic and microacupuncture because it is believed that the stimulations
haemorrhagic stroke (37% and 47% increase, resp.), number of the eye around the orbital margin can open the meridians,
of deaths (21% and 20% increase), and Disability Adjusted invigorate blood, stop pain, calm the “Shen,” and regulate
of Life Years (DALYs) lost (18% and 14% increase) [2]. Eye “Zang Fu” function [4].
acupuncture is a specialized and clinic approved acupuncture Standardized manipulation of eye acupuncture is various
treatment. It was invented by Doctor Jing-Shan Peng, the [5]. It could be the vertical insertion within the orbital
cavity, horizontal insertion outside the orbital cavity, prick- to Nursing and Allied Health Literature (CINAHL), The
ing acupuncture, double insertion, and successive insertion Chinese Biological Medicine Database (Sino Med), China
within and outside the orbital cavity. National Knowledge Infrastructure (CNKI), VIP Database,
Since it was invented, the eye acupuncture has been and Wan fang Database.
practiced in Liaoning University of Traditional Medicine for The search activity was ended in April 2014. The follow-
more than 40 years. Thousands of stroke patients received ing search terms were included: Ischemic stroke, Cerebral
this special treatment. Eye acupuncture has produced a infarction, Cerebral hemorrhage, Cerebrovascular accident
tremendous clinical significance. Today, eye acupuncture is (CVA), Eye acupuncture, Random; Chinese phrases “zhong
widely used in clinical treatment including: cerebrovascular feng,” “nao cu zhong,” “nao xue guan bing,” “ban shen bu sui,”
disease, pain, neurological disorders, and mental disease [6, “pian tan,” “nao geng si,” “nao geng se,” “nao chu xue,” “nao yi
7]. xue,” “nao xue shuan,” “nao shuan se,” “qiang xi xing geng si,”
There are about 400 trials related to the eye acupuncture “yan zhen,” and “sui ji.’’
stored in the database of China National Knowledge Internet
(CNKI). It seems that there is a large data of applications of
eye acupuncture treatment for stroke. The problem is that, 2.4. Data Extraction and Quality Assessment. The literature
similar to other effective TCM treatments, it still lacks com- searching (BZH, ZYY), study selection (ZZX, ZYY), and data
prehensive evaluation and system review. Thus, systematic extraction (BZH, ZYY) were conducted by three independent
review and meta-analysis of eye acupuncture treatment are authors. The extracted data include the name of author, title
necessary and will have a great significance for study in stroke of study, year of publication, study size, age and gender of the
related treatment and rehabilitation. participants, outcomes, adverse effects, prick depths of eye
acupuncture, and eye acupoints for each study. Disagreement
was resolved by discussion, and consensus was reached
2. Material and Methods through a third party (LCR).
2.1. Protocol and Registration. A protocol of this sys-
tematic review was published in “eye acupuncture ther- 2.5. Data Analysis. Rev Man 5.0 software was used for data
apy for stroke: a systematic review of randomized con- analysis. The effect estimates were presented as relative risk
trolled trials” (http://www.crd.york.ac.uk/prospero/display (RR) and mean difference (MD) with a 95% confidence
record.asp?ID=CRD42014009632#.VHqcSNJPgoE). interval. If a sufficient number in randomized trials were
identified, the subgroup analyses for the outcomes, such as
2.2. Inclusion Criteria. As interventions, randomized con- ADL, MRS, OHS, NIHSS, CSS, MMT, HAMD, MMSE, and
trolled trials (RCT) and quasi-randomized controlled trials WST, would be carried out.
(Quasi-RCT) of eye acupuncture were included in this study. Meta-analysis could be performed if the trials had a good
There was no limitation on language of publication or homogeneity on study design, participants, interventions,
publication type. controls, and outcome measures. Heterogeneity [8] between
According to the clinical criteria of the World Health studies could be investigated by 𝐼2 statistic which quantifies
Organization (WHO 1970), patients without limitations on inconsistency across studies. If an 𝐼2 was larger than 50%,
age or gender were included if they were diagnosed as stroke it could indicate the possibility of heterogeneity. Both fixed
patients. Patients were confirmed by purely clinical features effect model and random effect model would be used if there
or by the result of computed tomography (CT) or magnetic was a possibility of statistical heterogeneity among trials.
resonance imaging (MRI). Patients with ischemic as well as The fixed effect model would be used for meta-analysis, if
hemorrhagic stroke but not subarachnoid hemorrhage or 𝐼2 is less than 50%. The missing data could be obtained
subdural hematoma were considered for inclusion in the from the original trial authors. If a sufficient number of
review. randomized trials were identified, the sensitivity analyses
The interventions include eye acupuncture and combined would be performed to explore the influence of trial quality
treatments, such as eye acupuncture combining with western for effect estimates. The adequacy of generation of allocation
medicine treatment, herbal treatment, rehabilitation therapy, sequence, concealment of allocation, doubles blinding, and
or other alternative treatments. The controls could be western use of intention-to-treat (yes or no) were included as the
medicine treatment, herbal treatment, rehabilitation therapy, quality components of methodology.
or other alternative treatments. Trials would be excluded
if it related to any acupuncture treatment other than eye
acupuncture in order to eliminate the influence of different 3. Results
acupuncture methods.
3.1. Description of Studies. 16 randomized trials [9–24] were
included in this review. Five trials were reported as thesis [11,
2.3. Identification and Selection of Studies. The relevant arti- 13, 14, 19, 22], and the remaining 11 trials were published in
cles in the following databases were searched: Cochrane Chinese journals. A flow chart depicting the search process
stroke Group Trials Register, The Chinese Stroke Trials Reg- and study selection is shown in Figure 1. 16 RCTs and a total of
ister, The Chinese Acupuncture Trials Register, MEDLINE, 1120 stroke patients were involved in this review (69 patients
EMBASE, Alternative Medicine Database, Cumulative Index per trial).
82 citations were identified across the databases
Identical duplicated citation: 31
51 full-text papers assessed for eligibility

Excluded (n = 35)
(i) Reviews about eye
acupuncture: 4
(ii) Not RCT or quasi-RCT: 19
(iii) Inappropriate control group: 3
(iv) Trials related to any
acupuncture treatment other
than eye acupuncture in eye
acupuncture intervention or
in the control group: 6
(v) Obvious false data: 3
16 studies included in the systematic review
Figure 1: Flow chart of study selection.
The intervention time point for ischemic stroke and concealment nor blinding method was used in all trials.
hemorrhagic stroke in this study was varying from 1–3 days No follow-up document was provided. Protocols were not
to more than 6 months. available. The missing data in three trials [12, 15, 25] were
The content of intervention includes eye acupuncture, eye not available. Methodological quality has been summarized
acupuncture combined with western medicine, TCM herbal in Figure 2.
treatment, and rehabilitation. The control included western
medicine, TCM herbal treatment, and rehabilitation. 3.3. Effects of Interventions. Results of meta-analysis were
The outcomes were different. As the primary outcome, listed in Table 2 (estimate effect of included trials in meta-
CSS (Chinese Stroke Scale) was reported in 7 trials [9, 11, 13– analyses).
15, 20, 22]. Activities of Daily Living (ADL) were reported in
three trials [12, 17, 22]. HAMD (Hamilton Depression Scale) 3.3.1. Changes of CSS at the End of Treatment. The outcome
and WST (water swallow test) were reported, respectively, in of CSS at the end of the treatment was measured in 8 trials
two trials [16, 23]. The first defecation time and constipation [9, 11, 13–15, 20, 22] with 452 patients. When eye acupuncture
symptoms were evaluated in one trial [19]. SSS (Scandinavian is combined with western medicine compared to western
Stroke Scale) and FMA (Fugl-Meyer Scale) were assessed in medicine [9, 13–15], there was an obvious difference (MD
one trial [17]. MMSE (Mini-Mental State Examination) was −4.24, 95% CI −5.59 to −2.89 Fixed, 𝐼2 = 31% Fixed). One
reported in one trial [18]. The ranked data for effect judgment trial [11] compared the eye acupuncture combined with TCM
based on clinic neurological function deficit scale (NDS) herbal treatment to TCM herbal treatment, and there was a
was applied in one trial [10]. As secondary results, the level clear difference (MD −2.89, 95% CI −4.15 to −1.63). There was
change of ET and that of CGRP were reported in three trials a significant difference between eye acupuncture combined
[10, 13, 14, 24] and the level change of FIB [9] was reported with rehabilitation and western medicine versus rehabilita-
for pathological improvement. The change of CRP level was tion with western medicine [22] (MD −2.40, 95% CI −3.76
observed in one trial [15]. VEGF (vascular endothelial growth to −1.04). There was no significant difference between eye
factor) at the end of treatment was detected in one trial [25]. acupuncture combined with rehabilitation and rehabilitation
The characteristic of all included studies has been presented [20] (RR −2.40, 95% CI −4.87 to 0.07).
in Table 1.
3.3.2. Changes of ADL at the End of Treatment. The change
3.2. Methodological Quality. The study shows that the quality of ADL score was measured in 3 trials [12, 17, 22] with
of all included trials is poor. Five trials [10, 13–15, 22] used 207 patients. Two of these trials [17, 22] were collected on
random number table to allocate treatment. Three trials continuous variable with 140 patients. The data in the other
[11, 19, 21] were quasi-randomized. In these 3 trials, the trial [12] were not available. There was a significant difference
patients were allocated alternately according to the visiting when eye acupuncture was combined with rehabilitation
time point with the doctors in hospital. Nine trials did not and western medicine versus rehabilitation and western
describe the details of sequence generation. Neither adequate medicine [17] (MD 17.60, 95% CI 14.19 to 21.01). One trial [22]
4
Table 1: Characteristic of all included trials.

Sample size Intervention
Age Ischemic or Area of eye acupuncture Prick
Study ID Study type (T/C, Eye acupuncture Duration Outcomes
(yr, T/C) hemorrhagic Control intervention intervention depth
male/female) intervention
Basic treatment
T: 66 Major acupoints: upper
Wang et al. (1) Shuxuening injection
T: 60 (32/28) (42∼70) (1) Eye acupuncture jiao, lower jiao
(2008) RCT Ischemic (extract of Ginkgo) 14 days NA CSS; FIB
C: 60 (29/31) C: 64 (2) Basic treatment Minor acupoints: liver,
[9] (2) Citicoline Injection
(41∼70) kidney, spleen, and heart
(3) low-dose aspirin
Basic treatment
Major acupoints: upper
Zhou et al. (1) Sodium Ozagrel
T: 60 (46/16) (1) Eye acupuncture jiao, lower jiao Minor 2 mm along the NDS; ET;
(2011) RCT NA Ischemic injection 15 days
C: 60 (42/18) (2) Basic treatment acupoints: liver, kidney, cavity orbital CGRP
[10] (2) Citicoline injection
spleen, and heart
(3) Low-dose aspirin
(1) Eye acupuncture Major acupoints: upper
Liu (2010) T: 28
Q-RCT NA Ischemic (2) Buyang Huanwu Buyang Huanwu decoction 3 weeks jiao, lower jiao, spleen, NA CSS
[11] C: 28
Decoction and heart
T: (1) Eye acupuncture Major acupoints: upper
Pang (1) Rehabilitation training
T: 34 (40∼70) Ischemic and (2) Rehabilitation training jiao, lower jiao 9∼10.5 mm in
(2006) RCT (2) Basic treatment: >38 days ADL
C: 34 C: hemorrhagic (3) Basic treatment: Minor acupoints: liver, orbit
[12] medicine was not available
(40∼70) medicine was not available kidney, and heart
T: 51.31 ± 13.25 Basic treatment Major acupoints: upper
(40∼75) (1) Sodium Ozagrel jiao, lower jiao
Cui (2009) T: 8 (5/3) (1) Eye acupuncture 3 mm along the CSS; ET;
RCT C: Ischemic injection 2 weeks Minor acupoints: liver,
[13] C: 10 (6/4) (2) Basic treatment cavity orbital CGRP
51.59 ± 12.89 (2) Citicoline injection kidney, heart, spleen,
(40∼75) (3) Low-dose Aspirin and large intestine
T: 52.29 ± 14.89 Basic treatment Major acupoints: upper
(40∼75) (1) Sodium Ozagrel jiao, lower jiao CSS;
Li (2010) T: 23 (12/11) (1) Eye acupuncture 2 mm along the
RCT C: Ischemic injection 2 weeks Minor acupoints: liver, ET;
[14] C: 25 (13/12) (2) Basic treatment cavity orbital
52.46 ± 13.35 (2) Citicoline injection kidney, heart, spleen, CGRP
(40∼75) (3) Low-dose dose aspirin and large intestine
Main eye acupoints:
T: 63.24 Basic treatment
Wang et al. upper jiao, lower jiao
T: 45 (24/21) (40∼70) (1) Eye acupuncture (1) Shuxuening injection
(2007) RCT Ischemic 14 days Minor acupoints: liver, NA CSS; CRP
C: 45 (26/19) C: 64.98 (2) Basic treatment (2) Citicoline injection
[15] kidney, heart, spleen,
(43∼70) (3) Low-dose aspirin
and stomach
T:
Li and Wang (1) Eye acupuncture
T: 50 (42∼75) Basic treatment: medicine Major acupoints: upper
(2009) RCT Ischemic (2) Basic treatment: 2 weeks 7.5 mm in orbit WST
C: 50 C: was not available jiao
(42∼75)
Table 1: Continued.
T: (1) Rehabilitation training Major acupoints: upper
68.1 ± 8.2 (1) Eye acupuncture based on the Bobath jiao, lower jiao.
Chen et al.
T: 40 (24/16) (40∼80) (2) Rehabilitation training (2) Basic treatment Minor acupoints: liver, SSS; ADL;
(2007) RCT Ischemic 3 months NA
C: 40 (22/18) C: based on the Bobath (a) t-PA gallbladder, kidney, FMA
[17]
67.3 ± 11.1 (3) Basic treatment (b) Aspirin heart, spleen, and
(40∼80) (c) Mannitol injection middle jiao
Li (1) Eye acupuncture
T: 25 T: (50∼75) Basic treatment: medicine Major acupoints: upper
(2009) RCT Ischemic (2) Basic treatment: 2 weeks 7.5 mm in orbit MMSE
C: 25 C: (50∼75) was not available jiao, kidney, and spleen
(1) Eye acupuncture First
T: (2) Basic treatment: (1) Basic treatment: defecation
Xi
T: 30 (16/14) (35∼75) Ischemic and medicine was not available medicine was not available Major acupoints: lower time;
(2011) Q-RCT 7 days NA
C: 30 (18/12) C: hemorrhagic (3) Rehabilitation (2) Rehabilitation training: jiao, lung, and spleen constipation
[19]
(35∼75) training: methods were methods were not available symptoms
not available evaluation
T:
Jiang (2009) T: 30 (40∼70) Ischemic and (1) Eye acupuncture 2 mm along the
RCT Rehabilitation training 48 days jiao, lower jiao, kidney, CSS

[20] C: 30 C: hemorrhagic (2) Rehabilitation training cavity orbital
and liver
(40∼70)
Basic treatment
(1) 20% mannitol
(2) Cerebrolysin
(3) Huatuo Zaizao pill
T: 54 (4) Hemorrghagic: PAMBA
Ren and Lin jiao, lower jiao
T: 30 (21/9) (32∼78) Ischemic and (1) Eye acupuncture Nimodipine Nifedipine 2 mm along the Treatment
(2005) Q-RCT 30 days Minor acupoints: liver,
C: 28 (20/8) C: 53 hemorrhagic (2) Basic treatment (5) Ischemic: Low cavity orbital efficiency
[21] gallbladder, kidney, and
(37∼83) molecular dextran
heart
ATP Cytochrome C for
injection Dicoumarin
Nimodipine
Aspirin
Major acupoints: liver,
gallbladder, kidney, and
Basic treatment
heart.Minor acupoints:
Gao (2012) T: 30 (18/12) (1) Eye acupuncture (1) Aspirin or Clopidogrel 2 mm along the
RCT NA Ischemic 14 days upper jiao, lower jiao, ADL; CSS
[22] C: 30 (20/10) (2) Basic treatment Other medicines were not cavity orbital
heart, spleen, stomach,
available
large intestine, and
bladder
T:
Major acupoints: liver,
61.10 ± 10.12
Huang (2013) T: 80 (43/37) Ischemic and (1) Eye acupuncture middle jiao, heart Minor 2 mm along the
RCT (40∼76) Neurostan 8 weeks HAMD
[23] C: 76 (40/36) hemorrhagic (2) Neurostan acupoints: kidney, cavity orbital
C: 55.72 ± 9.02
spleen, and gallbladder
(40∼76)
5
6
Table 1: Continued.
Basic treatment: the
medicine might be
Xu et al. Xingding injection,
T: 34 (18/16) T: 62.5 ± 17.5 jiao, lower jiao Minor
(2006) RCT Ischemic Eye acupuncture compound danshen 22 days NA ET
C: 26 (16/10) C: 64.2 ± 7.7 acupoints: liver, kidney,
[24] injection, and
and heart
Deproteinized calf blood
injection
Basic treatment: the
medicine might be
Dong Deproteinized calf blood
T: 38 (20/18) T: 63.2 ± 12.5 Jiao, lower Jiao. Minor
(2009) RCT Ischemic Eye acupuncture injection, Shuxuening 7 days NA VEGF
C: 34 (18/16) C: 65.1 ± 8.6 acupoints: liver, kidney,
[25] injection (extract of
and heart
Ginkgo), Sanqi Panax
Notoginseng injection
Notes: (1) ADL: Activities of Daily Living. (2) CGRP: calcitonin gene related peptide. (3) CRP: C-reactive protein. (4) CSS: Chinese Stroke Scale. (5) ET: endothelin. (6) FIB: fibrinogen. (7) FMA: Fugl-Meyer
Scale. (8) HAMD: Hamilton Depression Scale. (9) MMSE: Mini-Mental State Examination. (10) SSS: Scandinavian Stroke Scale. (11) NDS: clinic neurological function deficit scale. (12) VEGF: vascular endothelial
growth factor.
Random sequence generation (selection bias)

Allocation concealment (selection bias)
Blinding of participants and personnel (performance bias)
Blinding of outcome assessment (detection bias)
Incomplete outcome data (attrition bias)
Selective reporting (reporting bias)
Other bias
0 25 50 75 100
(%)
Low risk of bias

Unclear risk of bias
High risk of bias
Figure 2: Methodological quality.
indicated that there was a significant difference between eye medicine versus rehabilitation and western medicine, there
acupuncture combined with western medicine treatment and was a significant difference in the constipation symptoms
western medicine treatment (MD 4.67, 95% CI 1.45 to 7.89). evaluation (MD −4.78, 95% CI −5.14 to −4.42) as well as the
first defecation time (MD −1.03, 95% CI −1.46 to −0.60).
3.3.3. Changes of SSS at the End of Treatment. The SSS
score at the end of treatment was applied in 1 trial [17] 3.3.9. Changes of NDS. The changes of NDS score at the end
with 80 patients. There was a significant difference between of treatment were checked in 1 trial [10] with 120 patients.
acupuncture combined rehabilitation and western medicine There was a significant difference between eye acupuncture
versus rehabilitation and western medicine (MD 12.41, 95% combined with western medicine and western medicine (RR
CI, 8.92 to 15.90). 1.08, 95% CI 0.93 to 126).
3.3.4. Changes of FMA Assessment at the End of Treatment. 3.3.10. Changes of ET Level at the End of Treatment. The
The FMA assessment at the end of treatment was applied changes of ET level at the end of treatment were checked in 4
in 1 trial [17] with 80 patients. When eye acupuncture was trials [10, 13, 14, 24] with 246 patients.
combined with rehabilitation and western medicine versus There was no significant difference between eye acupunc-
rehabilitation and western medicine, there was a significant ture combined with western medicine treatment and western
difference (MD 8.31, 95% CI, 3.15 to 13.47). medicine treatment in 2 trials [10, 14] (MD, 30.40, 𝐼2 = 100%,
95% CI −43.65 to 104.46 Random). There was no significant
3.3.5. Changes of HAMD Score at the End of Treatment. The difference between eye acupuncture combined with rehabil-
changes of HAMD at the end of treatment were observed in 1 itation and rehabilitation in 1 trial [13] (MD −10.71, 95% CI
trial [23] with 156 patients. There was no significant difference −28.9 to 6.67). There was a significant difference between eye
between eye acupuncture combined with western medicine acupuncture and western medicine treatment in 1 trial [24]
and western medicine (MD −0.82, 95% CI −1.87 to 0.23). (MD, −0.64, 95% CI −1.17 to −0.12).
3.3.6. Changes of MMSE at the End of Treatment. The changes
of MMSE at the end of treatment were observed in 1 trial [18] 3.3.11. Changes of CGRP Level at the End of Treatment.
with 50 patients. There was a significant difference between There was no significant difference between eye acupuncture
eye acupuncture combined with western medicine treatment combined with western medicine treatment and western
and western medicine treatment (MD 1.60, 95% CI 0.28 to medicine treatment (MD 1.48, 95% CI −5.31 to 8.27, 𝐼2 = 28%,
2.92). Fixed).
3.3.7. Changes of SWT at the End of Treatment. The changes 3.3.12. Changes of FIB Level at the End of Treatment. The
of SWT at the end of treatment were observed in 1 trial [16] changes of FIB level at the end of treatment were observed in
with 100 patients. There was a significant difference between 1 trial [9] with 120 patients. There was a significant difference
the eye acupuncture combined with western medicine and between eye acupuncture combined with western medicine
western medicine (RR 1.24, 95% CI 1.03 to 1.49). treatment and western medicine treatment (MD −0.72, 95%
CI, −1.09 to −0.35).
3.3.8. Changes of Constipation Symptoms and First Defecation
Time Evaluation at the End of Treatment. The first defecation 3.3.13. Changes of CRP Level at the End of Treatment. The
time and constipation symptoms at the end of treatment were changes of CRP level at the end of treatment were observed in
evaluated in 1 trial [19] with 60 patients. In the comparison of 1 trial [15] with 90 patients. There was a significant difference
eye acupuncture combined with rehabilitation and western between eye acupuncture combined with western medicine
Table 2: Estimated effect sizes of included trials in meta-analyses.
Trials Participants Estimate effects

(1) Changes of CSS scores for neurological assessment
(1.1) Eye acupuncture combined with western medicine versus western medicine
Wang et al. (2008) [9] 120 MD −5.56 [−7.15, −3.97]
Cui (2009) [13] 18 MD −3.23 [−9.14, 2.68]
Li (2010) [14] 48 MD −2.44 [−5.44, 0.56]
Wang et al. (2007) [15] 90 MD −3.84 [−5.35, −2.33]
Subtotal MD −4.24, 95% CI −5.59 to −2.89 𝐼2 = 31% fixed
(1.2) Eye acupuncture combined with TCM herbal treatment versus TCM herbal treatment
Liu (2010) [11] 56 MD −2.89 [−4.15, −1.63]
(1.3) Eye acupuncture combined with rehabilitation versus rehabilitation
Jiang (2009) [20] 60 RR −2.40 [−4.87, 0.07]
(1.4) Eye acupuncture combined with rehabilitation and western medicine versus rehabilitation
combined with western medicine
Gao (2012) [22] 60 MD −2.40 [−3.76, −1.04]
(2) Changes of ADL at the end of treatment
Chen et al. (2007) [17] 80 MD 17.60 [14.19, 21.01]
Gao (2012) [22] 60 MD 4.67 [1.45, 7.89]
(3) Changes of SSS score at the end of treatment
Chen et al. (2007) [17] 80 MD 12.41 [8.92, 15.90]
(4) Changes of FMA assessment at the end of treatment
Chen et al. (2007) [17] 80 MD 8.31 [3.15, 13.47]
(5) Changes of HAMD score at the end of treatment
Huang (2013) [23] 156 MD −0.82 [−1.87, 0.23]
(6) Changes of MMSE assessment at the end of treatment
Li (2009) [18] 50 MD 1.60 [0.28, 2.92]
(7) Changes of SWT assessment at the end of treatment
Li and Wang (2009) [16] 100 RR 1.24 [1.03, 1.49]
(8) Changes of NDS
Zhou et al. (2011) [10] 120 RR 1.08 [0.93, 1.26]
(9) Constipation symptoms evaluation at the end of treatment
Xi (2011) [19] 60 MD −4.78 [−5.14, −4.42]
(10) First defecation time at the end of treatment
Xi (2011) [19] 60 MD −1.03 [−1.46, −0.60]
Table 2: Continued.
Trials Participants Estimate effects
(11) Changes of ET level at the end of treatment
Zhou et al. (2011) [10] 120 MD −7.03 [−9.48, −5.12]
Li (2010) [14] 48 MD 68.27 [61.03, 75.51]
Subtotal 168 MD 30.40 95% CI −43.65 to 104.46 𝐼2 = 100% random
(11.2) Eye acupuncture combined with rehabilitation versus rehabilitation
Cui (2009) [13] 18 MD −10.71 [−28.09, 6.67]
(11.3) Eye acupuncture versus rehabilitation
Xu et al. (2006) [24] 60 MD −0.64 [−1.17, −0.12]
(12) Changes of CGRP level at the end of treatment
Zhou et al. (2011) [10] 120 MD 5.67 [4.03, 7.31]
Li (2010) [14] 48 MD 1.48 [−5.31, 8.27]
Subtotal 168 MD 1.48 95% CI −5.31 to 8.27 𝐼2 = 28% fixed
(13) Changes of FIB level at the end of treatment
Wang et al. (2008) [9] 120 MD −0.72 [−1.09, −0.35]
(14) Changes of CRP level at the end of treatment
Wang et al. (2007) [15] 90 MD −5.86 [−7.54, −4.18]
(15) Changes of VEGF level at the end of treatment
(15.1) Eye acupuncture versus western medicine
Dong (2009) [25] 60 MD 0.02 [−0.49, 0.53]
treatment and western medicine treatment (MD −5.86, 95% 4. Discussion

CI, −7.54 to −4.18).
The focal points in this study are the safety and effectiveness
of eye acupuncture for stroke. The study demonstrated that
3.3.14. Changes of VEGF Level at the End of Treatment. The eye acupuncture is a safe and effective treatment for stroke
changes of VEGF level at the end of treatment were observed patients on symptoms alleviation and the dependency in the
in 1 trial [25] with 60 patients. There was no significant results of CSS, SSS, ADL, FMA, MMSE, HAMD, WST, and
difference between eye acupuncture and western medicine first defecation time as well as the biochemistries tests (CRP,
(MD 0.02, 95% CI, −0.49 to 0.53). ET, VEGF, and CGRP).
Ignoring the methodological quality of included tri- However, there were several limitations of this review.
als, the results showed some effect in independency and The quality of the included studies was poor because there
symptom alleviation. As eye acupuncture is combined with were a mass of trials either having high or unclear risk of
western medicine versus western medicine, effects appeared bias. 13% trials of random sequence generation results were
in the outcomes of CSS, ADL, FIB, SWT, CRP, and FIB. of high risk and 53% trials were unclear. One trial was high
Eye acupuncture combined with TCM herbal treatment risk and the rest were unclear in blinding and the same result
showed more effectiveness than TCM herbal treatment in appeared again in allocation concealment. No trials about
the outcome of CSS. The outcomes of ADL have showed adverse events or death were mentioned so it was impossible
superiority of eye acupuncture combined with rehabilitation to get any information about safety and no trials have had the
compared to rehabilitation. The outcomes of ADL, SSS, FMA, follow-up observation either. There were some therapeutic
constipation symptoms, and first defecation time were more effects, but the outcomes did not focus on commonly used
effective in eye acupuncture combined with rehabilitation evaluation standards.
and western medicine as compared to rehabilitation and There were eight areas and thirteen points for eye
western medicine. acupuncture, but it was noticeable that the location of eye
acupoints is different in trials according to the mentioned
3.4. Adverse Event. No adverse events or side effects have intervention methods. We wish that the locations of eye
been reported during or after the eye acupuncture treatment acupoints could be unified according to Standardized Manip-
according to the trials. ulations of Eye Acupuncture [5]. Furthermore, trials of
eye acupuncture therapy should follow the Standards for cerebral infarction patients,” Shanghai Journal of Acupuncture
Reporting Interventions in Clinical Trials of Acupuncture and Moxibustion, vol. 27, no. 3, pp. 5–7, 2008.
(STRICTA) [26] to confirm the effect in stroke and facilitate [10] H. F. Zhou, J. Wang, M. B. Zhang et al., “Effect of eye acupunc-
a meta-analysis. It has been recommended that primary ture on endothelin and calcitonin in patients in the acute stage
outcome measures for stroke should be at the level of of cerebral infarction,” Shanghai Journal of Acupuncture and
Activities of Daily Living and the outcome should be assessed Moxibustion, vol. 30, no. 10, pp. 651–653, 2011.
at 6 months [27]. [11] C. Liu, The combined treatment of eye acupuncture and
There was no data indicating adverse events or death. Buyanghuanwu Tang for stroke at the recovery period, clinical
observation [M.S. thesis], Liaoning University of Traditional
But considering the position of needling, prick depths, sense
Chinese Medicine, Shenyang, China, 2010.
of fear that patients might confront, and other potential
[12] L. L. Pang, “Clinical observation of 68 cases in the combined
risks for stroke, the author strongly suggests safety evaluation
treatment of eye acupuncture and rehabilitation for stroke,” Jilin
and psychology evaluation should be carried out for eye Journal of Traditional Chinese Medicine, vol. 26, no. 5, pp. 49–50,
acupuncture. 2006.
[13] N. Cui, The gene influence of endothelin and calcitonin for
Conflict of Interests ischemic stroke patients with treatment of eye acupuncture [M.S.
thesis], Liaoning University of Traditional Chinese Medicine,
The authors declare that they have no conflict of interests in Shenyang, China, 2009.
the research. [14] S. Q. Li, The influence of endothelin and calcitonin for ischemic
stroke patients with treatment of eye acupuncture [M.S. the-
Authors’ Contribution sis], Liaoning University of Traditional Chinese Medicine,
Shenyang, China, 2010.
Zeng-Hua Bai and Zhi-Xing Zhang have equally contributed [15] P. Q. Wang, J. L. Li, and J. Wang, “Influences of eye needles in
to this work. acute brain stem dead patient of nerve dainage-function and
in blood serum C-response protein level,” Journal of Clinical
Acknowledgments Acupuncture and Moxibustion, vol. 23, no. 12, pp. 23–25, 2007.
[16] Z. Li and J. Wang, “Eye acupuncture therapy in treating
This work was supported by the Natural Science Foundation acute cerebral infarction complicated by dysphagia clinical
of Liaoning Province of China (201102147) and the funding observation,” Journal of Practical Traditional Chinese Internal
was from TCM Scholars Apprenticeship Project and Special Medicine, vol. 23, no. 9, pp. 89–90, 2009.
Program of Chinese Medicine of the National Basic Research [17] Y. F. Chen, M. Mao, and X. Chen, “Eye acupuncture combined
Program of China (973 Program, no. 2012CB518503). with exercise therapy for the treatment of limb dyskinesia due to
cerebral infarction: a report of 40 cases,” Journal of New Chinese
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http://dx.doi.org/10.1155/2015/646872
Research Article
Effect of Acupuncture on Functional Connectivity of
Anterior Cingulate Cortex for Bell’s Palsy Patients with
Different Clinical Duration
Hongli Wu,1,2 Hongxing Kan,1 Chuanfu Li,2 Kyungmo Park,3 Yifang Zhu,2
Abdalla Z. Mohamed,3 Chunsheng Xu,2 Yuanyuan Wu,2 Wei Zhang,2 and Jun Yang2
1
Medical Information Engineering, Anhui University of Chinese Medicine, Hefei, Anhui 230012, China
2
Laboratory of Digital Medical Imaging, Medical Imaging Center, The First Affiliated Hospital of Anhui University of Chinese Medicine,
Hefei, Anhui 230031, China
3
Department of Biomedical Engineering, Kyung Hee University, Yongin 446701, Republic of Korea
Correspondence should be addressed to Hongxing Kan; ffdkhx@ahtcm.edu.cn and Jun Yang; yangzyun@yahoo.cn
Received 5 December 2014; Revised 31 March 2015; Accepted 16 April 2015
Copyright © 2015 Hongli Wu et al. This is an open access article distributed under the Creative Commons Attribution License,
Acupuncture is widely used in the treatment of Bell’s palsy (BP) in many countries, but its underlying physiological mechanism
remained controversial. In order to explore the potential mechanism, changes of functional connectivity (FC) of anterior cingulate
gyrus (ACC) were investigated. We collected 20 healthy (control group) participants and 28 BP patients with different clinical
duration accepted resting state functional MRI (rfMRI) scans before and after acupuncture, respectively. The FC of ACC before
and after acupuncture was compared with paired 𝑡-test and the detailed results are presented in the paper. Our results showed
that effects of the acupuncture on FC were closely related to clinical duration in patients with BP, which suggested that brain
response to acupuncture was closely connected with the status of brain functional connectivity and implied that acupuncture plays
a homeostatic role in the BP treatment.
1. Introduction mechanism of acupuncture for treating BP remain contro-

versial and need further investigation. By now, only a few
Bell’s palsy is a unilateral idiopathic and mostly transient studies have investigated the effect of acupuncture on resting
facial paralysis resulting from dysfunction of the cranial state functional magnetic resonance imaging (rfMRI) FC
nerve VII (the facial nerve), with a pure peripheral deef- of BP patients. Our preliminary research concerning the
ferentation [1]. Previous researches have demonstrated that instant effect of acupuncture in BP treatment [5] reported
the cortical reorganization may complement the recovery negative activation in the early stage and positive activation
from facial nerve palsy with the change of functional con- in the later stage of BP. Besides, another investigation [6]
nectivity (FC), mainly demonstrated as disruption at the early of our group showed that changes in FC of the primary
stage and enforcement at the later stage [2] in the areas related somatosensory cortex (SI) induced by acupuncture varied
to error detection, sensorimotor integration, motor integra- with clinical durations of BP, mainly displayed as decreased
tion, and control. The FC [2], defined as the temporal correla- connectivity in the early stage but increased connectivity
tion between spatially remote neurophysiological events, has in the later stage. Furthermore, our rfMRI FC studies [7]
become a significant method for studying neuroplasticity to reported that FC of ACC showed positive correlation with
detect changes during cortical reorganization. the duration of BP, which suggested that ACC may play a
Acupuncture, as an alternative and complementary thera- crucial role in the process of cortical reorganization during
peutic intervention, is playing an increasingly important role the recovery from BP. Based on these critical conclusions,
in treating BP [3, 4]. However, effectiveness and underlying in order to probe the effect of acupuncture on the FC of BP
Patient group Healthy
HBS
HBS > 1 HBS = 1

HBS
Duration < 14 d Duration > 14 d
Early group Late group Recovered group Healthy group

18 cases 21 cases 19 cases 20 cases
9 males 14 males 10 males 11 males
19–70 years old 19–70 years old 19–63 years old 23–54 years old
10 left facial pareses 12 left facial pareses 7 left facial pareses
Figure 1: Healthy group and four subgroups of patient group classified based on House-Brackmann score (HBS) and disease duration.
patients and further reveal the underlying role of acupuncture the Human Research Committee of the First Affiliated Hospi-
in the cortical reorganization in the recovery process of BP, we tal of Anhui University of Traditional Chinese Medicine (see
investigated longitudinal changes of FC (before versus after Figure 1).
acupuncture) of bilateral ACC for BP patients with different
clinical duration. 2.2. fMRI Data Acquisition. The experiment was performed
The hypothesis of the present research was that acupunc- in the MRI room of the Medical Imaging Center, the First
ture may have different effect on the FC of ACC for patients Affiliated Hospital of Anhui University of TCM. The Siemens
with different clinical duration during the recovery of BP. Symphony 1.5 T MRI whole body scanner and standard head
coil were used. All subjects were instructed to lie down with
2. Materials and Methods eyes closed and to stay awake. All lights in the scanning room
were turned off to avoid unwanted visual stimulation.
2.1. Subjects. All subjects recruited in this study were right- Eight sequences were scanned: (1) pilot images; (2) T2-
handed with no histories of drug abuse and no mental, central weighted images to rule out any disease of the brain; (3) T1-
nervous system or other serious disease. The subjects were weighted 2D anatomical images with the axial position par-
divided into two groups: the patient group (28 cases, totally allel to the AC-PC line; the images include 36 slices that cov-
58 times MRI scanning) and the healthy control group (20 ered the whole brain. T1-weighted spin-echo sequence was
cases, totally 20 times MRI scanning). used, with TR/TE = 500/12 ms, FOV = 230 mm × 230 mm,
The patient groups, including patients with left and right slice thickness/interval = 3.0 mm/0.75 mm, and resolution =
unilateral facial paresis, were further divided into three 192 × 144; (4) resting-state fMRI before acupuncture,
subgroups based on the disease duration and HBS scores namely, Run 1; the EPI BOLD sequence with TR/TE/FA =
(House-Brackmann facial nerve grading system, 1 = normal 3000 ms/30 ms/90∘ was used and FOV = 192 mm × 192 mm
facial movement, and 6 = no movement, scored by an and resolution = 64 × 64; (5) resting-state fMRI with
experienced acupuncturist with no prior knowledge about the same parameters during acupuncture, namely, Run
the data results). The 3 subgroups (stages) were the early 2; (6) resting-state fMRI with the same parameters after
group (18 cases, 9 males, 19–70 years old; duration < 14 d, acupuncture, namely, Run 3; (7) task-state acupuncture fMRI:
HBS > 1; 10 left facial pareses), the late group (21 cases, 14
the scanning direction and the number of slices were the
males, 19–70 years old; duration > 14 d, HBS > 1; 12 left facial
same as those of the resting-state fMRI, with TR/TE/FA =
pareses), and the recovered group (19 cases, 10 males, 19–
4000 ms/50 ms/90∘ , namely, Run 4; (8) T1-weighted 3D
63 years old; HBS = 1; 7 left facial pareses). Among the 28
anatomical images: the sagittal position was taken, and
subjects of the patient group with different clinical duration,
total of 176 slices were scanned which covered the whole
4 undertook MRI scanning only once, 18 undertook it twice,
brain. The spoiled gradient echo sequence was used, with
and 6 undertook it thrice. Manual acupuncture was applied
TR/TE/FA = 2100 mm/3.93 mm/13∘ , FOV of 250 × 250 mm,
to all patients thrice a week semi-individually at acupoints
slice thickness/spacing = 1.0 mm/0.5 mm and resolution of
chosen by acupuncturists based on the individual symptoms
in the course of acupuncture treatment. The acupoint Hegu 256 × 256. It took about 60 minutes to complete all of the
(LI4) was selected in present experiment for all subjects since data acquisition. fMRI paradigms are shown in Figure 2.
LI4 is usually selected as the main acupoints in the clinical
treatment of BP. All healthy subjects (20 cases, 11 males, 23– 2.3. Extraction of the Region of Interest. Bilateral ACC was
54 years old) were either college students or the workers extracted as region of interest (ROI) for FC analysis. The
in the hospital. All subjects signed informed consent forms datasets were from 37 healthy volunteers with the task of
before participating in the experiment in accordance with mouth movement. The paradigm for motor task lasted 400
Needle-in Needle-out
Run 1 Run 2 Run 3 Run 4
Resting-state fMRI Resting-state fMRI Resting-state fMRI Task-state fMRI
before acupuncture during acupuncture after acupuncture
200 points, 10 min 200 points, 10 min 200 points, 10 min 160 points, 10 min
Rotate needles
10 s every 2 min
Figure 2: fMRI paradigms of the experiment.
seconds, which consists of 40-time task of opening and Tool) and melodic to compensate for any head movements
shutting mouth (mean time 7.8 ± 1.6 s) separated by 2-second during the scan. The functional images were then coregis-
duration. The volunteers were trained to open or shut mouth tered to the high-resolution anatomical images reconstructed
when seeing the word (outward/protrude) shown randomly by Freesurfer. Afterwards, the functional images were reg-
on the display before scanning and were instructed to lie istrated to standard MNI152 space (Montreal Neurological
down on the scanning bed and keep their body static. Institute) using FNIRT and FLIRT (affine transformation
The experiment was completed in the Department of with FMRIB’s Linear Image Registration Tool). Functional
Biomedical Engineering of Kyung Hee University of South data were smoothed using a Gaussian kernel of FWHM
Korea. The Philips Achieva 3.0 T MRI whole body scanner 6 mm; and band-pass filter (0.007 HZ < 𝑓 < 0.1 HZ) was
and 8-channel head coil were used. A total of 3 sequences also performed to reduce the effect of low-frequency drift and
were scanned, which were (1) 2D structural image: TR/TE of high-frequency noise. Then, individual data of right-sided
2000 ms/35 ms, voxel size of 2.785 mm × 2.875 mm × 4 mm, facial palsy patients were flipped along the 𝑦-axis so that all
slice/volume of 34/180, and matrix 80 × 80; (2) EPI-BOLD: data could be processed unilaterally. The individual 4D data
TR/TE of 2000 ms/35 ms, voxel size of 2.785 mm × 2.875 mm was then used for further group statistics and connectivity
× 4 mm, slice/volume of 34/200, and matrix of 80 × 80; (3) analysis.
T1-Weighted 3D structural image: TR/TE of 2000 ms/35 ms,
slice/gap of 1.0 mm/1.0 mm, and matrix of 256 × 256. 2.6. Functional Connectivity Analysis. In our research, to
To investigate the changed FC of bilateral ACC with compute the FC of ACC, the temporal signal series of
related brain areas, the ROIs were extracted from the statistic cerebrospinal fluid, white matter was extracted firstly. Then,
activation maps obtained from motor task experiment. The based on linear regression analysis, variances including 6
maximum point of activation strength of bilateral ACC, along parameters obtained by rigid body correction of head motion
with its 33 neighbors, was selected as ROIs (a sphere with and the signal of cerebral spinal fluid and white matter were
radius as 4 mm and voxel size 2 × 2 × 2 mm), as shown in removed. Finally, individual statistical maps were obtained
Figure 1 of [7]. based on the general linear model for further group analysis.
2.4. Paradigms of the Experiment. The acupuncture in the 2.7. Group Analysis. Standard space parameters of individual
experiment was executed by a professional acupuncturist. subjects were imported to a high level analysis with FLAME
Resting-state fMRI data before acupuncture (Run 1) lasts for (FMRIB’s Local Analysis of Mixed Effects). With the aim of
10 min (200 points). Then, the needle was inserted into the investigating the effects of acupuncture on FC of BP patients
acupoints of LI4 on the contralateral hand of the paralyzed with different clinical durations, we conducted intergroup
face and rotated to achieve De-Qi sensation. The second fMRI analysis through paired 𝑡-test before and after acupuncture
data (Run 2) was collected including 200 points for 10 min, for each group. The individual data with head movement
during which the needle was rotated bidirectionally for 10 s more than 2 mm or 2∘ were excluded before group analysis.
every 2 min. Then the needle was pulled out and the third The significance threshold for FEAT was set at 𝑧 = 2.3
fMRI data after acupuncture (Run 3) including 200 points and 𝑃 = 0.01. All results were then corrected using cluster
was obtained. (based on theory of Gaussian Random Field, GRF) to obtain
the activation maps. The results of intergroup analysis were
2.5. Data Preprocessing. Data analysis was performed using corrected using Monte-Carlo simulations with 𝑃 = 0.01,
the FSL (Oxford Centre for Functional MRI of the Brain’s 𝛼 = 0.05, and cluster size = 68.
(FMRIB’s) Software Library), Freesurfer, and AFNI. The
preprocessing was applied as follows. Anatomical images 3. Results
were reconstructed using Freesurfer recon-all and then tilt
correction was done for functional and anatomical images There were no significant difference among subjects’ age
using 3drefit and fslorient. Nonbrain removal was done using distributing and sample size of four groups. To address
mri watershed for anatomical images and BET for func- the significant differences in functional connectivity changes
tional images. Motion correction was done using MCFLIRT among different groups, the results of group analysis for each
(Motion Correction using FMRIB’s Linear Image Regression group were showed as follows.
Table 1: Group analysis of areas changed FC with right ACC after acupuncture for patient with Bell’s palsy in the early group.
Coordinate (MNI)
Region (BA) Side BA 𝑍 value Cluster size
Peak 𝑥 (mm) Peak 𝑦 (mm) Peak 𝑧 (mm)
Superior frontal gyrus R 8 32 28 56 −4.151 143
Middle frontal gyrus R 6 26 18 62 −3.643 126
Middle frontal gyrus R 10 38 30 46 −3.515 77
BA: Brodmann area; L: left; R: right; the threshold was set at 𝑃 ≦ 0.01; 𝛼 ≦ 0.05 (corrected with Monte-Carlo method).
Table 2: Group analysis of areas changed FC with right ACC after acupuncture for patient with Bell’s palsy in the latter group.
Coordinate (MNI)
Region (BA) Side BA 𝑍 value Cluster size
Peak 𝑥 (mm) Peak 𝑦 (mm) Peak 𝑧 (mm)
Superior temporal gyrus R 22 58 2 −4 3.945 130
Insula R 22 46 6 −4 3.633 102
Superior temporal gyrus R 41 46 −34 12 3.806 88
Putamen R 28 8 2 3.626 68
BA: Brodmann area; L: left; R: right; the threshold was set at 𝑃 ≦ 0.01; 𝛼 ≦ 0.05 (corrected with Monte-Carlo method).
3.1. The Healthy Control Group. Paired 𝑡-test was done before 4. Discussions
and after acupuncture to find out significant changes induced
by acupuncture effect on FC of bilateral ACC of the healthy In this study, in order to assess the role of acupuncture during
group. In the healthy group, after being corrected with the recovery of BP, we investigated the FC changes of ACC
Monte-Carlo method (𝑃 = 0.01, 𝛼 = 0.05 and cluster size = induced by acupuncture for patients with different clinical
duration. The results suggest that the acupuncture effects on
68), nostatistical significant differences were observed for
the FC varied with clinical duration.
bilateral ACC before and after acupuncture. The results imply
that acupuncture has no significant effect on FC of bilateral
ACC in healthy subjects. 4.1. Overall Acupuncture Effect on ACC Connectivity in Bell’s
Palsy Patients with Different Clinical Durations. As well
known, one of the most important characteristics of BP is
3.2. The Early Group. Different results were observed for the the damaged efferent nerve (without afferent nerve) and the
early group. In the early group, after being corrected with consequent impaired facial motor function of the affected
Monte-Carlo method (𝑃 = 0.01, 𝛼 = 0.05, and cluster size = side of the face. Therefore, the sensory feedback of the
68), significant changed FC were found after acupuncture. acute reduction of facial motor performance due to BP will
For the left ACC (ipsilateral to Bell’s palsy), no remarkable be detected by brain, thus causing increased processing in
changes of FC were found. Significant decreased connectivity brain areas responsible for the monitoring and integration of
of the right ACC (contralateral to BP) after acupuncture were somatosensory and motor information. Previous functional
observed in right superior frontal gyrus (SFG, BA 8), right neuroimaging studies have shown that the ACC is important
middle frontal gyrus (MFG, BA 6), and right middle frontal in error detection and performance-monitoring functions,
gyrus (MFG, BA 10) (as shown in Figure 3 and Table 1). including executive function, response selection, and conflict
monitoring [8].
3.3. The Latter Group. For the latter group, after being The present results indicated that no changes were found
corrected with Monte-Carlo method (𝑃 = 0.01, 𝛼 = 0.05, for the FC status of bilateral ACC for both the healthy and
and cluster size = 68), FC of ACC was also changed after the recovered groups after acupuncture. Presently, researches
acupuncture. For the left ACC (ipsilateral to Bell’s palsy), no show that the therapeutic principles of acupuncture are not
significant changes of FC were found. Significant increased through relieving local condition of the diseased area but in
the way of reestablishing the balance of the internal milieu
FC of the right ACC were observed in right superior temporal
(involving Ying/Yang, the Five Elements, and the Zhong-Fus)
gyrus (STG, BA 22), right insula (BA 22), right superior tem-
[9]. Therefore, for healthy subjects and the recovered group,
poral gyrus (STG, BA 41), and right putamen (see Figure 4
their homeostatic can be considered to stay in/in return to
and Table 2). a balance state; thus no significant effect on FC of ACC was
observed after acupuncture.
3.4. The Recovered Group. Similar with the healthy group, in Acupuncture-induced FC changes of the contralateral
the recovered group, the intergroup analysis results before ACC were observed for both the early and the later groups. As
and after acupuncture with Monte-Carlo correction (𝑃 = we know, one of the most remarkable features of the human
0.01, 𝛼 = 0.05, and cluster size = 68) showed no significant brain is its ability to adapt to new situations and to new
functional connectivity changes of bilateral ACC. information. Changes in the functional network connectivity
L R
MFG (BA 6)
SFG (BA 8)
MFG (BA 10)
−4 +4
Figure 3: Changed functional connectivity of right ACC after acupuncture for patients with Bell’s palsy at early stage. 𝑃 ≤ 0.01, 𝛼 ≤ 0.05,
corrected with Monte-Carlo method. BA: Brodmann area; SFG: superior frontal gyrus; MFG: right middle frontal gyrus; MFG: middle frontal
gyrus.
status, which reflect the process of cortical reorganization of the contralateral ACC and thus the FC status changed due
different brain areas, may give expression to this feature. It to various adjustment and compensatory mechanism. No
is suggested that [1] the functional reorganization caused by changes of the FC status of the ipsilateral ACC may indicate
transient peripheral deefferentation, which can be interpreted that the brain areas related to the healthy side remain in a
as the compensatory effect of brain to the impaired motor good condition.
performance of BP patients, happened after Bell’s palsy. As
a result of this process, the connectivity status of the brain 4.2. Changed Functional Connectivity of Sensorimotor Related
changed and thus led to a different acupuncture response Brain Areas. Compared with the Pre-Acu, decreased FC was
of brain. Generally speaking, compared with the healthy found in sensorimotor related brain areas including SFG (BA
group, FC changes in the early and the late group imply the 8) and MFG (BA 6, BA 10) after acupuncture. BA 8 and BA
different acupuncture responses resulting from the changed 6 are well known as the advanced motor center in planning,
FC status of BP patients. The results here are consistent integration, and execution of motor function. The difficulty
with our previous research concerning the instant effect of in movement of the paralyzed facial muscle in patients with
acupuncture in BP treatment, which reported the conclusion BP may elicit FC changes of these motor related areas.
that the brain responses to acupuncture differ at different Studies concerning the BP suggested that the impaired motor
pathological statuses and probably depend on the brain function (without a lesion in the brain) might initially lead
functional status [1]. to a disrupted connectivity within the cortical facial motor
Another remarkable characteristic of our results is that network, with a subsequent reorganization supporting func-
the acupuncture-induced changes of the intrahemispheric FC tional recovery [1, 2]. Our research concerning acupuncture-
were found only for the contralateral ACC but not for the induced FC changes of SI also reported decreased connectiv-
ipsilateral ACC for all groups. Previous investigations have ity in the early stage and increased connectivity in the later
consistently reported that the changed brain responses to stage [1]. Therefore, hypoactivation of motor related areas
acupuncture were mainly contralateral to the palsy area [2]. after acupuncture may result from disruption FC in the early
An fMRI study on BP [2] also revealed a significant acutely stage of Bell’s palsy.
disrupted but unaltered interhemispheric connectivity of MI Another sensorimotor correlated brain area with changed
and other parts of the facial motor network at early stage FC was observed in putamen at the latter group. While
of the palsy, followed by a return toward normal during it has been concluded that the putamen has no specific
the course of recovery. Their results are in accordance with specialization, it works with many other motor related
ours. This can be explained since motor commands of paretic structures to control many types of motor skills, including
side are blocked in the acute stage of BP; then the error motor learning, motor performance and tasks [10], and motor
in the process of command execution was feed backed to preparation [11]. In addition, the putamen contains high
L R
STG (BA 41)
Insula
STG
R putamen
−4 +4
Figure 4: Changed functional connectivity of right ACC after acupuncture for patients with Bell’s palsy at later stage. 𝑃 ≤ 0.01, 𝛼 ≤ 0.05,
corrected with Monte-Carlo method. BA: Brodmann area; STG: superior temporal gyrus; STG: superior temporal gyrus.
levels of opioid receptors and is considered to be involved 4.3. Changed Functional Connectivity of Homeostatic Related
in sensory and emotional components of pain. Meanwhile, Network. Changed connectivity between the contralateral
fMRI research of patients with PD received acupuncture ACC and insular cortex induced by acupuncture is also
treatment demonstrated that the activated putamen was an interesting finding in our research. As well known, the
correlated with enhanced motor function of patients. Based insular cortex is involved in a wide range of functions includ-
on comprehensive analysis, increased FC in putamen may ing motor control and homeostatic regulation [12–15]. As
result from the cortical reorganization in motor correlated reported in previous neuroimaging researches, the ACC and
brain areas which were caused by the loss of facial motor insular cortex play an important role in the network termed as
control in BP patients. the “homeostatic afferent processing network” [16]. The net-
In addition, increased FC was observed in superior work represents all aspects of the physiological condition of
temporal gurus (STG, BA 22, and BA 41), which is well known the body and meanwhile provides crucial sensory input that
as the primary auditory cortex and is involved in auditory and is essential for maintaining homeostasis. The fMRI research
language processing. Changed FC in temporal lobe reflects in functional diarrhea reported that acupuncture brings
brain’s potential compensatory mechanism because of facial functional connectivity changes to the homeostatic afferent
motor difficulty in BP patients. Besides, changed FC in STG processing network only in patients but not in the healthy
(BA 22) may also occur as cortical reorganization result from group because of functional abnormality of the network in
the difficulty in flexible pronunciation of those patients with functional diarrhea patients. The results were in accordance
serious facial paralysis. with ours since the changed FC induced by acupuncture
within the homeostatic afferent processing network in BP [2] C. M. Klingner, G. F. Volk, S. Brodoehl, H. P. Burmeister, O.
patients may suggest the unbalanced functional state result W. Witte, and O. Guntinas-Lichius, “Time course of cortical
from the Bell’s palsy. All results imply the homeostatic plasticity after facial nerve palsy: a single-case study,” Neurore-
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of acupuncture and moxibustion to treat Bell’s palsy according
self-recovery rate of Bell’s palsy and there was no control
to different stages: design and protocol,” Contemporary Clinical
group in our experiment, the results might just reflect the self- Trials, vol. 30, no. 4, pp. 347–353, 2009.
recovery of the disorder instead of the acupuncture effect. [4] L.-A. Liu, Y.-H. Zhu, Q.-Q. Li, and Z.-L. Yu, “Comparison of
efficacy and the safety evaluation of electroacupuncture with
5. Conclusion and Limitation different waveforms for peripheral facial paralysis patients,”
World Journal of Acupuncture-Moxibustion, vol. 23, pp. 17–21,
The paper introduces the influence of acupuncture on FC 2013.
of ACC for BP patients with different clinical duration and [5] C. Li, J. Yang, J. Sun et al., “Brain responses to acupuncture are
our results indicate that different changes were observed. probably dependent on the brain functional status,” Evidence-
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than one functional neural system. The deactivation in the Article ID 175278, 14 pages, 2013.
early group and positive activation in the latter group kept [6] R. J. Morecraft, K. S. Stilwell-Morecraft, and W. R. Rossing,
“The motor cortex and facial expression: new insights from
peace with the disrupted connectivity in the acute stage and
neuroscience,” Neurologist, vol. 10, no. 5, pp. 235–249, 2004.
subsequent reorganization during the recovery of the BP. This
[7] S. Hu, Y. Y. Wu, C. F. Li et al., “Increasing functional connectivity
synchronization suggested the dependence of acupuncture of the anterior cingulate cortex during the course of recovery
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homeostatic role of acupuncture in BP treatment. Moreover, [8] S. Zhang and C.-S. R. Li, “A neural measure of behavioral
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induced by acupuncture also reflected homeostatic regulation level-dependent activity in the precuneus,” NeuroImage, vol. 49,
role of acupuncture. no. 2, pp. 1911–1918, 2010.
Present results indicate that acupuncture might at least [9] L. Leung, “Neurophysiological basis of acupuncture-induced
partly contribute to the functional recovery of BP arising analgesia—an updated review,” Journal of Acupuncture and
from the cortical reorganization by changing the FC within Meridian Studies, vol. 5, no. 6, pp. 261–270, 2012.
the somatosensory motor network and other related areas. [10] M. R. DeLong, G. E. Alexander, A. P. Georgopoulos, M. D.
The evidence provided in this research for the effectiveness Crutcher, S. J. Mitchell, and R. T. Richardson, “Role of basal
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of acupuncture in treating BP is limited because there was 4, pp. 235–244, 1984.
no blank control group due to ethics restriction. Therefore,
[11] G. E. Alexander and M. D. Crutcher, “Preparation for move-
we cannot exclude the possibility that the results might just ment: neural representations of intended direction in three
reflect the self-recovery of Bell’s palsy. The paper may bring motor areas of the monkey,” Journal of Neurophysiology, vol. 64,
to light the underlying mechanism of acupuncture during the no. 1, pp. 133–150, 1990.
recovery of BP, although further researches are still needed. [12] K. Xu, H. Liu, H. Li et al., “Amplitude of low-frequency
fluctuations in bipolar disorder: a resting state fMRI study,”
Conflict of Interests Journal of Affective Disorders, vol. 152–154, no. 1, pp. 237–242,
2014.
The authors declare that there is no conflict of interests [13] I. Mutschler, B. Wieckhorst, S. Kowalevski et al., “Functional
regarding the publication of this paper. organization of the human anterior insular cortex,” Neuro-
science Letters, vol. 457, no. 2, pp. 66–70, 2009.
[14] J. A. Avery, W. C. Drevets, S. E. Moseman, J. Bodurka, J. C.
Acknowledgments Barcalow, and W. K. Simmons, “Major depressive disorder is
associated with abnormal interoceptive activity and functional
The study was supported by the National Key Basic Research connectivity in the insula,” Biological Psychiatry, vol. 76, no. 3,
and Development Program (973) (2010CB530500), the pp. 258–266, 2014.
National Natural Science Foundation of China (81202768), [15] E. G. Duerden, M. Arsalidou, M. Lee, and M. J. Taylor, “Later-
the Natural Science Foundation of Anhui Traditional Chi- alization of affective processing in the insula,” NeuroImage, vol.
nese Medicine University (2014zr019), and the Youth Foun- 78, pp. 159–175, 2013.
dation of Anhui Traditional Chinese Medicine University [16] S. Zhou, F. Zeng, J. Liu et al., “Influence of acupuncture stim-
(2014qn030, 2011qn022). ulation on cerebral network in functional diarrhea,” Evidence-
Based Complementary and Alternative Medicine, vol. 2013,
Article ID 975769, 9 pages, 2013.
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http://dx.doi.org/10.1155/2015/158012
Research Article
The Study of Dynamic Characteristic of Acupoints
Based on the Primary Dysmenorrhea Patients with
the Tenderness Reflection on Diji (SP 8)
Sheng Chen,1 Yanhuan Miao,2 Yinan Nan,3 Yanping Wang,2 Qi Zhao,1

Enhui He,1 Yini Sun,1 and Jiping Zhao1
1
Acupuncture and Moxibustion Department, Dongzhimen Hospital, Beijing University of Chinese Medicine,
No. 5 Haiyuncang, Dongcheng District, Beijing 100010, China
2
College of Acupuncture-Moxibustion and Tuina, Beijing University of Chinese Medicine, No. 13 of the North 3rd East Road,
Chaoyang District, Beijing 100029, China
3
International Medical Center, China-Japan Friendship Hospital, No. 2 Yinghua East Street, Chaoyang District, Beijing 100029, China
Correspondence should be addressed to Jiping Zhao; zjp7883@sina.com
Received 19 December 2014; Revised 3 March 2015; Accepted 3 March 2015
Copyright © 2015 Sheng Chen et al. This is an open access article distributed under the Creative Commons Attribution License,
In TCM theory, acupoint is not a fixed point but a portal with dynamic characteristics where the channel qi enters and flows out. The
dynamic characteristics have been verified primarily by detecting the tenderness reaction on Diji (SP 8) in primary dysmenorrhea
patients. In this study, finger pressing and algometer were applied in Diji (SP 8) area of participants in menstrual period and
nonmenstrual period, respectively, to detect the tenderness occurrence rate, the VAS score of the tenderest point, the tenderness
threshold of the tenderest point, and the location of the tenderest point. The result suggests that the acupoint may not be a fixed
location but a point in a dynamic state within a certain range in time and space varying with different physiological and pathological
status.
1. Background 2010. The standardization of the acupoint location has to

some extent normalized clinical operation and promoted
Acupoints are specific locations where qi and blood of the dissemination and development of the acupuncture and
meridians and Zang-fu organs infuse and also where diseases moxibustion worldwide. However what catches our attention
are reflected and the acupuncture needles are applied [1]. is that while promoting the standard of the acupoint location,
Therefore, acupoint is the essential factor in acupuncture parts of the clinical practitioners stick too much to the
diagnosis and treatment. Each acupoint has its own name, fixed location of the acupoint and ignore the importance
location, specificity, function, and so forth, while location and necessity of searching and seeking acupoint along the
is considered the most essential and fundamental. Without meridian, leading to dissatisfactory clinical effect.
accurate and precise location, it is likely neither to examine According to the description in the Yellow Emperor’s Inner
the reaction of the acupoint in pathological condition, nor Canon, the classic of Traditional Chinese Medicine, acupoint is
to find the point with therapeutic effects. Because of this, a portal where the channel qi enters and flows out and not a
the standardization of the acupoint location is considered fixed point attaching to the skin, vessel, muscle, tendon, and
as the key in the history of Chinese acupuncture and bone. Therefore, acupoint is not isolated or static structure
moxibustion standardization. Since the official promulgation in the body. It is related to the movement of channel qi. It
of the location of acupoints (national standard) in 1990, is not only able to reflect the changes of qi in channel, but
updated standards of the acupoint locations continued to also able to be used for adjusting the channel qi, showing
be introduced one after the other in the years 2006 and dynamic characteristics. Whereas most of the researches on
Table 1: Comparison ages between observation group and control group (unit: age).
Age
Group Number of cases
Smallest value Largest value M (QR)
Observation group 30 21 34 26 (5)
Control group 30 23 32 25 (2)
𝑍 −0.88
𝑃 0.378
dynamic performance of acupoints have been theoretical, (1) The first one is women with lower abdominal pain
clinical research is lacking. that begins somewhere between several hours before and a
Diji (SP 8) is the most important and commonly used few hours after the onset of the menstrual bleeding, usually
point for the treatment of dysmenorrhea. According to the persisting up to 2-3 days; (2) the pain is characteristically
theory of traditional Chinese medicine (TCM), Diji (SP 8) is colicky or dull and located in the midline of the lower
the Xi-cleft point of the Spleen meridian of Foot Taiyin, where abdomen but may extend to both lower quadrants, the lum-
the meridian qi accumulates deeply and is suitable for treating bar area, and the thighs; (3) the pain is frequently associated
acute pain and blood disease. Primary dysmenorrhea, a with symptoms including diarrhea, nausea, vomiting, fatigue,
medical condition of cramping pain in the lower abdomen light-headedness, headache, dizziness, and, rarely, syncope
occurring before and during menstruation, is just ascribed and fever; (4) the symptoms are more or less reproducible
to acute pain and blood disease in TCM. Therefore, this from one menstrual period to the other; (5) type B ultrasonic
research focuses on patients with primary dysmenorrhea to examination and gynecological examination exclude the
observe the changes in tenderness in Diji (SP 8) in different organic pathological changes in the reproductive organs.
physiological and pathological states, so as to explore the
dynamic characteristic of acupoint and to provide clinical (2) Inclusion Criteria. These criteria include the following: (1)
data for the study of acupoint dynamism from the clinical patients who fulfill the diagnostic standard of the primary
perspective. dysmenorrhea; (2) patients of ages between 18 and 35 years;
(3) patients who have never given birth; (4) patients who
have disease duration ≥6 months; (5) patients who have
2. Materials and Methods
regular menstrual cycle (28 ± 7) d; (6) patients who have
2.1. Setting and Participants. 30 patients with primary dys- abdominal pain which occurs 48 hrs within the onset of men-
menorrhea were recruited as the observation group between struation; (7) patients with COX Dysmenorrhea Symptom
April and December of 2013 in Dongzhimen Hospital affiliated Scale (CMSS) [3] total score ≥8; (8) patients with VAS score of
to Beijing University of Chinese Medicine. 30 healthy female the abdomen pain ≥40 during the attack of dysmenorrhea; (9)
volunteers from the Beijing University of Chinese Medicine patients who have no participation in any other medication
were recruited as the control group during the same period or modality clinical trials; (10) patients who signed informed
of time. consent.
As some patients with primary dysmenorrhea may
resolve or be relieved spontaneously after giving birth, all (3) Exclusion Criteria. These criteria include the following:
the participants included were nulliparous, so as to reduce (1) patients with life threatening disorders, such as cardio-
selection bias. In the observation group, the oldest participant vascular, liver, kidney, hematopoietic system disorders, and
was 34 years old, the youngest was 21 years old, and the mental diseases; (2) patients who have skin problem on and
mean of their age was 26 ± 5 years. In the control group, near Diji (SP 8), such as soft tissue damage, ulceration, scar,
the oldest was 32 years old, the youngest was 32 years old, and skin calluses; (3) patients who received other related
the smallest age was 23 years, and the mean of their age was treatments within a month or intake of pain killers, sedatives,
25 ± 2 years. After the statistical analysis of the distribution and hormone drugs within 2 weeks; (4) patients who are
of age and disease duration between the control group and physically weak or judged not suitable to participate in this
the observation group, the differences were not statistically research by researchers.
significant. See details in Table 1.
2.2.2. Control Group
2.2. Diagnostic Criteria, Inclusion Criteria, and
(1) Inclusion Criteria. These criteria include the following:
Exclusion Criteria
(1) healthy women with no abnormalities in the physical
2.2.1. Observation Group examination within the recent semester; (2) patients of ages
between 18 and 35 years; (3) patients who have never given
(1) Diagnosis Criteria. Referring to the Canadian Department birth; (4) patients who have no history of dysmenorrhea in
of Gynecology and Obstetrics Association in 2005 primary the past; (5) patients who have mostly regular menstrual cycle
dysmenorrhea clinical guideline [2], standards are as follows. (28 ± 7) d; (6) patients who signed informed consent.
Yinlingquan
(SP 9)
3 B-cun
Diji (SP 8)
Figure 2: Algometer.
Figure 1: Standard position of Diji (SP 8).

the position. The point with highest VAS scores was the
tenderest point; if there is no tenderness upon pressure, the
(2) Exclusion Criteria. These criteria include the following: result was just recorded with no VAS detection.
(1) patients suffering from frequent lower abdominal pain
of unknown reason; (2) patients with skin problems on and 2.3.3. Detecting Pain Threshold of the Tenderest Point. After
near Diji (SP 8), such as soft tissue damage, ulceration, scar, the VAS assessment, the participants were instructed to rest
and skin calluses; (3) patients who received other related for 10 minutes. Then, the pain threshold of the tenderest
treatments within a month or intake of pain killers, sedatives, point was detected using an algometer (National Patent
and hormone drugs within 2 weeks; (4) patients with mental number: ZL200520142236.5; Product Publication number:
disorders or judged not suitable to participate in this research CN2862954; Manufacturer: Institute of Orthopedics and
by researchers. Traumatology Affiliated to Chinese Academy of Traditional
Chinese Medicine Science; Place of Production: Beijing)
2.3. Tenderness Detection on Diji (SP 8). In order to guarantee (Figure 2).
the quality of the study, every segment of the operation was Firstly set the tester to zero. Then put the probe tip
performed by the same researcher, who had received training (0.5 cm in diameter) of the tester vertically onto the mark
over six months, to ensure the standardization and unity of point. Apply pressure gradually and evenly downward (the
the operation. maximum pressure should not exceed 600 kpa for avoiding
tissue damage caused by excessive force). Once the partici-
2.3.1. Detecting Point. Diji (SP 8) on both sides of the legs and pant reports pain or a radiating pain was elicited, then the
their surrounding areas of a total of 60 participants in both algometer was removed and the data on the tester screen was
the observation group and control group was detected. recorded as the pain threshold value. Such a procedure was
conducted on both sides of the Diji (SP 8) area, with the left
The participants were instructed to lie supine with legs
one coming first.
straightened in a relaxed manner while fully exposing the
parts below the knees.
2.3.4. Measure the Location of the Tenderest Point. The
Diji (SP 8) was located referring to WHO standard
distance between the center of the tenderest point and the
acupuncture point locations in the Western Pacific Region [4],
standard position of Diji (SP 8) was measured with a soft tape
which is “on the tibial aspect of the leg, posterior to the medial
measure and then recorded.
border of the tibia, 3 B-cun inferior to SP9” (Figure 1). Mark
this point as the standard position of Diji (SP 8).
2.3.5. Detecting Time Point
2.3.2. Detecting Tenderness with VAS [5]. Starting from the
(1) For the observation group, detection of time point
standard position of Diji (SP 8), the researcher pressed
occurred during the first day or second day following
spirally with the tip of the thumb pulp in a circular area
the onset of dysmenorrhea and the seventh day after
2 cm long in radius. The intensity of the pressing force was
menstruation (nonmenstrual period).
consistent and even to the level of muscle. The tenderness
was recognized when pain, soreness, or distension sensation (2) For the control group, detection of time point took
was expressed through the immediate and fleeting reactions place the first or second day following menstrual
of participants’ eyes or words. onset and the seventh day after menstruation (non-
When tenderness reaction appeared, the participants menstrual period).
were instructed to face the reverse side of the VAS card
without graduation and then move the cursor to the position 2.4. Statistical Analysis. SPSS17.0 statistical software was used
that best represented the pain intensity. The researcher facing for analysis. Count data were tested using 𝑥2 test. One-
the side with calibration recorded VAS scores and marked way ANOVA was adopted for sets of normally distributed
Pressing pain 60
70
60
50
50 19
31 38
40 53 40
30
VAS
20 41 30
10 29
22
7 20
0
non-menstrual
menstrual period
Control group,
Observation group,
Observation group,
menstrual period
Control group,
non-menstrual
10
period
period 0
menstrual period
non-menstrual period
Control group-Non,
Observation group,
menstrual period
Control group,
menstrual period
Observation group,
Negetive
Positive
Figure 3: Comparison of the TOR of each group during menstrual

and nonmenstrual period.
Figure 4: Comparison of VAS score between menstrual period and
nonmenstrual period in two groups (mm).
data which went through paired comparison using S-L-D
method. 𝑡-test was used for the data from two groups and 3.2. Comparison of VAS Score of the Tenderest Point in Diji
the data were expressed by the mean plus or minus standard (SP 8) Area. Using nonparametric test, we compared VAS
deviation. Skewed data were tested using the nonparametric score between groups in menstrual period and nonmenstrual
Wilcoxon test and expressed in M (QR), that is, the median period (corrected value 𝑃󸀠 = 0.00833). VAS score of the
(interquartile range). All statistical tests were tested and observation group in the menstrual period was significantly
verified using the two-sided test. 𝑃 ≤ 0.05 was considered higher than that in the nonmenstrual period, while there was
statistically significant. no significant difference between two periods in the control
group. In menstrual period, VAS score of the observation
group was significantly higher than that of the control
3. Results group, 𝑃 < 𝑃󸀠 . In nonmenstrual period, VAS score of the
observation group was higher than that of the control group,
3.1. Comparisons of Tenderness Occurrence Rate in Diji (SP 8)
while there was no significant difference, 𝑃 > 𝑃󸀠 (Figure 4,
Area. In observation group, there were 5 one-side tenderness
Table 3).
cases and 1 pain-free case; the rest of the cases presented with
tenderness on both sides in Diji (SP 8) area during acute
onset period. Total TOR was 88.3%. During nonmenstrual 3.3. Comparison of Tenderness Threshold Value of the Ten-
derest Point in Diji (SP 8) Area. We used one-way ANOVA
period, there were 8 one-side tenderness cases and 7 pain-free
to analyze two groups’ tenderness threshold value of the
cases, and the rest presented with tenderness on both sides.
tenderest point in Diji (SP 8) area in menstrual and nonmen-
Total TOR was 63.3%. In control group, there were 11 one- strual period. The results showed 𝐹 = 4.983, 𝑃 = 0.003 <
side tenderness cases and 9 pain-free cases, and the rest of the 0.05, indicating that there was difference in threshold value
cases presented with tenderness on both sides in Diji (SP 8) between two groups in menstrual or nonmenstrual period.
area during menstrual period. Total TOR was 51.7%. During S-L-D method was used for further analysis. Tenderness
nonmenstrual period, there were 5 one-side tenderness cases threshold value in the menstrual period of the observation
and 18 pain-free cases, and the rest presented with tenderness group was significantly lower than that in the nonmenstrual
on both sides. Total TOR was 31.7%. period, 𝑃 < 𝑃󸀠 , while there was no significant difference
We used Chi-squared test to compare the TOR between between two periods in the control group. In menstrual
two groups and two menstrual periods (corrected value period, tenderness threshold value of the observation group
𝑃󸀠 = 0.00833). In the observation group, the TOR during was significantly lower than that of the control group, 𝑃 < 𝑃󸀠 .
menstrual period was significantly higher than that in the In nonmenstrual period, tenderness threshold value of the
nonmenstrual period, 𝑃 < 𝑃󸀠 . In the control group, the observation group was lower than that of the control group,
TOR during menstrual period was also significantly higher while there was no significant difference, 𝑃 > 𝑃󸀠 (Figure 5,
than that in the nonmenstrual period, 𝑃 < 𝑃󸀠 . During Table 4).
nonmenstrual period, the TOR of the observation group was
also significantly higher than that of the control group, 𝑃 < 𝑃󸀠 3.4. Location of the Tenderest Point in Diji (SP 8) Area. Our
(Figure 3, Table 2). results showed overlaps between the tenderest point and
Table 2: Comparison of the TOR in Diji (SP 8) area between menstrual and nonmenstrual period.
Group Menstrual period Nonmenstrual period 𝑥2 𝑃

Positive 53 38
Tenderness
Observation group Negative 7 22 10.231 0.001
TOR 88.3% 63.3%∗
Positive 31 19
Tenderness
Control group Negative 29 41 4.937 0.026
TOR 51.7%∗ 31.7%#
𝑥2 19.206 12.063
𝑃 0.000 0.001
Note: corrected value 𝑃󸀠 = 0.00833. ∗ Compared with observation group in menstrual period, 𝑃 < 𝑃󸀠 . # Compared with observation group in nonmenstrual
period, 𝑃 < 𝑃󸀠 .
Table 3: Comparison of VAS score between menstrual and nonmenstrual period in two groups (mm).
Group Menstrual period Nonmenstrual period 𝑍 𝑃

Number of effective values 53 38
Minimum value 10 5
Observation group Maximum value 60 40 −2.646 0.0081
M (QR) 30 (20) 22.5 (25)∗
Mean rank 88.76 66.12
Minimum value 5 5
Control group Maximum value 40 40 −0.182 0.856
M (QR) 20 (15)∗ 20 (20)
Mean rank 54.89 57.50
𝑍 −3.705 −0.780
𝑃 0.000 0.436
Note: corrected value 𝑃󸀠 = 0.00833. ∗ Compared with observation group in menstrual period, 𝑃 < 𝑃󸀠 .
2.00 standard point in observation group during menstrual period

and nonmenstrual period. Overlap rate was 22.6% and 28.9%,
respectively. We measured the distance between the tenderest
Pressing pain’s threshold value
1.50 point and the standard point and came up with the following
conclusion: if we set standard point of Diji (SP 8) as datum
point, distribution range of the tenderest point in observation
1.00 group was 0.565–0.903 cm in menstrual period and 0.515–
0.974 cm in nonmenstrual period, respectively (Figure 6,
Table 5).
0.50
4. Discussions
0.00
The dynamic characteristic of the acupuncture point is one of
the hot topics in the acupuncture research. Previous study has
menstrual period
non-menstrual period
Control group-Non,
Observation group,
menstrual period
Control group,
menstrual period
Observation group,
shown that acupuncture points are of dimensional structure

located in the interstice within the skin, vessel, muscle,
sinew, bone, and even viscera, rather than fixed points. Their
location may be influenced by several factors such as different
physiological changes and pathological conditions of the
Zang-fu organs and channels and the external environment
Figure 5: Tenderness threshold value in Diji (SP 8) area in mens- and individual variety. Therefore, acupuncture points possess
trual and nonmenstrual period (kPa). the individualized and dynamic characteristic [6].
Table 4: Tenderness threshold value in Diji (SP 8) area in menstrual and nonmenstrual period (kPa).
Group Menstrual period Nonmenstrual period Mean difference 𝑃

Minimum value 0.01 0.1
Observation group −0.251 0.001
Maximum value 1.2 1.6
𝑥±𝑠 0.497 ± 0.040 0.748 ± 0.375∗
Minimum value 0.1 0.06
Control group −0.027 0.801
Maximum value 1.61 1.62
𝑥±𝑠 0.724 ± 0.385∗ 0.751 ± 0.468
Mean difference 0.227 −0.003
𝑃 0.006 0.979
Note: corrected value 𝑃󸀠 = 0.00833. ∗ Compared with observation group in menstrual period, 𝑃 < 𝑃󸀠 .
Table 5: Distance between the tenderest point and standard point in Diji (SP 8) area (cm).
Number of Overlap rate with Minimum value Maximum value M (QR) 95% CI
effective values standard point
Menstrual period 53 22.6% 0 3 0.7 0.6 0.565–0.903
Observation group
Nonmenstrual period 38 28.9% 0 2.6 0.7 (1) 0.515–0.974
There are four reasons for applying the pressing examina- group were higher than that in nonmenstrual period
tion on Diji (SP 8) of the patients who suffered from primary and the menstrual period in the control group. The
dysmenorrheal to study the dynamism of acupuncture points tenderness threshold of Diji (SP 8) in the menstrual
on reflecting disease in this research. First, according to period of observation group was lower than that in
the channel and acupuncture point theory in TCM, Diji the nonmenstrual period of observation group and
(SP 8) is the Xi-cleft point of the Spleen meridian of Foot the menstrual period in the control group. The result
Taiyin. Xi-cleft points are where the meridian qi accumulates showed that there exist dynamic characteristics in Diji
deeply and are indicated for the acute and pain disease of (SP 8) in the tender reaction in both the physiological
the respective Zang-fu organs and meridians, while the Xi- and pathological conditions, including the different
cleft points of the Yin meridians are also indicated for blood stages in the pathological condition. The tenderness
diseases [7]. Primary dysmenorrhea, a medical condition of reaction was more likely to occur and more intensive
cramping pain in the lower abdomen occurring just before and sensitive in the menstrual period of the primary
and during menstruation, is ascribed to acute pain and dysmenorrhea patient.
blood disease in TCM. Therefore, Diji (SP 8) is the most
(2) There is no statistical difference in the tenderness
important and commonly used point for the treatment of
occurrence rates, VAS score, and tenderness thresh-
dysmenorrhea [8–11]. Second, the acupuncture points relate
old in both the menstrual period and nonmenstrual
closely to the internal Zang-fu organs through the pathway
period in the control group. It indicates that there
of the meridians; thereby the condition of the diseased Zang-
is no remarkable change in the tenderness reaction
fu organs will be reflected on the acupuncture point through
on Diji (SP 8) in the physiological state and the
the transmission of the meridians [12]. Third, though there
alteration of the physiological rhythm. Comparing
are various examination methods and techniques for the
the data in the nonmenstrual period of the test
reflection effect of acupuncture point, such as detecting the
and control group, the tenderness occurrence rate of
electric currency and electrical resistance of the point [13–
observation group is higher than that of the control
17] and the infrared thermal imaging technique [18, 19], the
group, but there is no statistical difference between
most commonly used, convenient, and consensus method is
the two groups in VAS value and the tenderness
detecting the tenderness and pain threshold [20–22]. Fourth,
threshold. The results might be considered as fol-
there are few researches and reports on reflecting effect of Diji
lows: in the nonmenstrual period, the dysmenorrhea
(SP 8) on the dysmenorrhea.
patients were still in the pathological states of blood
Diji (SP 8) of the 30 patients with primary dysmenorrhea
deficiency failing to nourish the uterus or blood
and 30 healthy women was palpated by hand and detected
stasis blocking the meridian in the uterine, which
by algometer. The result of the research and the concerning
makes Diji (SP 8) become more sensitive to pressing;
issues are discussed as follows.
however the severity was not intensive. Or it might
(1) The tenderness occurrence rates and the VAS score be that the patients enrolled were not enough to
of Diji (SP 8) in menstrual period of the observation show the statistical difference. From an anatomical
Observation group, non-menstrual period Observation group, menstrual period

3 3
2 2
1 1
0 0
−1 −1
−2 −2
−3 −3
−3 −2 −1 0 1 2 3 −3 −2 −1 0 1 2 3
VAS value VAS value

Diji (SP 8) 21–40 Diji (SP 8) 21–40
0–20 41–60 0–20 41–60
(a) (b)
Figure 6: Location of the tenderest point in Diji (SP 8) area (cm).
perspective, there are parts of the saphenous nerve in pathological reactions such as pain, tenderness, and
shallow layer of Diji (SP 8) area and sympathetic nerve other changes on the acupuncture point [26]. Zeng et
governing the myometrium contraction coming from al. put forward the viewpoint that, in the pathological
the same nerve segments. Accordingly, the same state, the surface reflection area of acupuncture varies
situation also occurs between the tibial nerve in deep with the condition of the disease; it increases when
layer and parasympathetic nerve controlling the sense disease gets worse, decreases when disease gets better,
of uterus. It can thus be seen that Diji (SP 8) has and disappears when disease is cured [27, 28]. As to
a close relationship with uterus in anatomy [23]. the enlargement of the surface areas around acupoint
However, the results still possibly indicate that Diji reflecting diseases, Yu et al. believe that it relates to the
(SP 8) might reflect the pathological condition in facilitation and sensitization of the spinal cord caused
the menstrual period of the dysmenorrhea patients, by visceral disorders, where the information coming
which also validate the viewpoint that pathological from the body surface and viscera are assembled [29].
reaction in meridian and acupuncture points relates According to the theory of “painful locality taken
to timing [24]. as an acupoint,” the tenderest point in Diji (SP 8)
(3) This research found out that, in Diji (SP 8) area, area might be the veracious point of Diji (SP 8)
the tenderest points were divergent from the stan- in the pathological state and it might also be the
dardized location of Diji (SP 8) in the majority most effective point for treatment. We have found in
of dysmenorrheal patients, which indicates that the clinic that, in the acute stage of dysmenorrhea, the
location of Diji (SP 8) in pathological state is different part between Sanyinjiao (SP 6) and Yinlingquan (SP
from the standardized location. Acupuncture point 9) on the pathway of Spleen meridian is the main
is both the reflective point of diseases and where reaction region. Tender, sore, or distending points,
the needles and moxibustion are applied for the especially distinctive around the Sanyinjiao (SP 6),
treatment of disease in the meridian and acupunc- Diji (SP 8), and Yinlingquan (SP 9) area, will be found
ture point theory [25]. Wang illuminates the process when palpating along the meridian. Needling on the
acupuncture reflecting disease. Acupuncture point is tender point will bring instinctive effect of relieving
where both the pathological factors and meridian the pain. Of course, large scale clinical trial is required
qi exist when patient suffered from a disease. If the to confirm our clinical observation.
meridian qi fails to dispel the pathological qi out of the (4) There have been 2 Chinese national standards of
body in time, the pathological factors accumulate in acupuncture location (1990 and 2006) and Stan-
the acupuncture point; consequently, there might be dard Acupuncture Nomenclature (second edition) was
published in 1993 [30–34]. The standardization of Conflict of Interests

acupuncture point location helps to standardize the
needle manipulation and also promote the spreading The authors declare that there is no conflict of interests
of acupuncture worldwide. In standardized acupunc- regarding the publication of this paper.
ture point location, a vertical and horizontal coor-
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Acupoint Sensitization, Acupuncture Analgesia, Acupucnture On Visceral Functional Disorders and Its Mechanism

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Evidence-Based Complementary and Alternative Medicine

Acupoint Sensitization, Acupuncture Analgesia,

Effects of Deep Electroacupuncture Stimulation at “Huantiao” (GB 30) on Expression of

Function of Nucleus Ventralis Posterior Lateralis Thalami in Acupoint Sensitization Phenomena,

Effect of Repeated Electroacupuncture Intervention on Hippocampal ERK and p38MAPK Signaling in

Correspondence should be addressed to Xiaochun Yu; yuxc@mail.cintcm.ac.cn

Received 14 May 2015; Accepted 21 May 2015

Correspondence should be addressed to Jiang Zhu; jzhjzh@263.net

Received 22 August 2014; Revised 9 November 2014; Accepted 19 November 2014

Academic Editor: Xinyan Gao

Allocated to Deqi Allocated to no Deqi

One treatment for 30 minutes

Evaluation of results at the end of treatment

Received 11 August 2014; Revised 18 September 2014; Accepted 3 October 2014

Academic Editor: Jian Kong

1. Introduction visceral sensation, and psychosocial factors have all been

Assessed for eligibility

Patients who accepted

Treatment group (n = 30) Control group (n = 30)

Figure 1: Flow chart of study participation.

Table 1: General characteristics of patients with FD prior to participation.

Items Groups 𝑁 Baseline After treatment Difference Improvement rate 𝑃 value

Table 3: Dyspeptic symptom sum score at baseline and during follow-up.

IND Groups 𝑁 Baseline Follow-up Difference Improvement 𝑃 value

Items Baseline (𝑛 = 8) After treatment (𝑛 = 8) Healthy adults (𝑛 = 10)

Received 18 December 2014; Revised 16 February 2015; Accepted 30 April 2015

Academic Editor: Zhixiu Lin

1. Introduction Vascular dementia (VaD) refers to cognitive impairment

Acupuncture, a primary therapeutic method in tradi-

2.1. Inclusion Criteria for Considering Acupoint

Random sequence generation

Blinding of participants and personnel

Blinding of outcome assessment

Incomplete outcome data

Figure 2: Whole view on the study quality of the 185 CCTs.

Fengchi (GB 20)

Shuigou (GV 26)

Shenting (GV 24)

Zusanli (ST 36)

Benshen (GB 13)

Shenshu (BL 23)

Fenglong (ST 40)

Dazhui (GV 14)

Xuanzhong (GB 39)

Fengfu (GV 16)

Quchi (LI 11)

Figure 3: The 20 most frequent acupoints and their frequencies.

Number Acupoint combination Frequency Support (%)

Eight conﬂuent acupoints

Lower He-Sea point

Eight conﬂuent acupoints

Lower He-Sea point

(a) Network structure of acupoints for the treatment of VaD

BL23 GV20 PC6

(b) 19-core network

Correspondence should be addressed to Tieming Ma; matieming999@sohu.com

Received 10 December 2014; Revised 30 January 2015; Accepted 5 February 2015

Academic Editor: Haifa Qiao